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RESEARCH I I iTFF SYSTEM

KrosFlo Research IIi TFF System Product Information and Operating

Instructions

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TRADEMARKS AND WARRANTY

ii KrosFlo® RESEARCH IIi TFF SYSTEM • PRODUCT INFORMATION AND OPERATING INSTRUCTIONS

Spectrum’s KrosFlo® Research IIi TFF System meets strict quality control standards and is warranted against

defects in material and workmanship for a period of two (2) years from date of shipment.

The information contained herein is believed to be accurate and is offered in good faith for the convenience of

the user. PRODUCTS ARE FURNISHED UPON THE CONDITION THAT THE USER ASSUMES ALL RISKS

AND LIABILITIES AND THAT NEITHER THE SELLER NOR MANUFACTURER SHALL BE LIABLE FOR ANY

LOSS OR DAMAGE, DIRECT OR CONSEQUENTIAL, ARISING FROM THE USE OF THESE PRODUCTS.

Spectrum®, KrosFlo®, MicroKros®, MidiKros® and MiniKros® are registered trademarks of Spectrum

Laboratories, Inc. Microsoft®, Windows® and Excel® are registered trademarks of Microsoft Corporation. C-

Flex® and Pharmapure® are registered trademarks of Saint-Gobain. MasterFlex® is a registered trademark of

Thermo Fisher Scientific. Trademarks bearing the ® symbol in this publication are registered in the U.S. and in

other countries.

This document copyright © 2012 by Spectrum Laboratories, Inc. All rights reserved. Reproduction prohibited

except by permission of the copyright owner.

1 KrosFlo® RESEARCH IIi TFF SYSTEM • PRODUCT INFORMATION AND OPERATING INSTRUCTIONS

TABLE OF CONTENTS

Table of Contents

System Overview and Operation1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3

2. Safety Precautions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3

3. Specifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5

4. System Configuration and Major Components . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6

4.1 KrosFlo® Research IIi Pump Drive & Pump Head . . . . . . . . . . . . . . . . . . . . . . . . . 6

4.2 KrosFlo® Research IIi Pressure Monitor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6

4.3 Flowpath Kit . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6

4.4 Starter Kit . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6

5. System Options and Parts List . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6

6. Materials of Construction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7

7. KrosFlo Research IIi Pump/Pressure Monitor Set Up and Operation. . . . . . . . . . . . . . 8

7.1 Installation and Setup . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8

7.2 Operation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9

7.2.1 Turning on the System. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9

7.2.2 Control Panel. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10

7.2.3 Home Screen . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10

7.2.4 Setup Screen. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11

A. Pressure Calibration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11

B. Alarm Settings. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12

C. Tubing Calibration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12

D. Hardware Setup . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14

E. Diagnostics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15

7.2.5 Troubleshooting and Error Messages. . . . . . . . . . . . . . . . . . . . . . . . . . . . 15

8. KrosFlo Research IIi Pump Head Set Up and Operation . . . . . . . . . . . . . . . . . . . . . . 18

8.1 Safety Precautions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18

8.2 Specifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19

8.3 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20

8.4 Installation and Removal . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20

8.5 Tubing Loading and Unloading . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21

8.6 Multi-Channel Mounting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22

8.7 Maintenance and Cleaning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22

9. KF Comm Software . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23

9.1 License. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23

9.2 Installation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 4

TABLE OF CONTENTS (continued)


9.3 Instructions for Use. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 24

9.3.1 Opening File . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . page 24

9.3.2 Data Collection Tools . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25

9.3.3 Navigating the Worksheets and Charts . . . . . . . . . . . . . . . . . . . . . . . . . 26

9.3.4 Compliancy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 29

10. TFF Filtration using Spectrum HF Modules . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30

10.1 Basic Concepts of Tangential Flow Filtration (TFF) . . . . . . . . . . . . . . . . . . . . . . 30

10.1.1 Dead-end Filtration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30

10.1.2 Tangential Flow Filtration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30

10.2 Assembling the Flowpath . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 32

10.2.1 Constant Volume Mode . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 32

10.2.2 Batch Mode . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39

10.3 Integrity Testing. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39

10.3.1 Leak Test. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39

10.3.2 Wetting the HF Membrane Module . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39

10.3.3 Pressure Hold Test . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 40

10.4 Modes of Operation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 41

10.4.1 Batch Concentration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 41

10.4.2 Batch Clarification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 41

10.4.3 Constant Volume . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 41

10.4.4 Diafiltration. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 42

10.5 Operating the System. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 43

10.5.1 Instructions for Batch Concentration / Clarification . . . . . . . . . . . . . . . . 43

10.5.2 Instructions for Constant Volume & Diafiltration . . . . . . . . . . . . . . . . . . . 44

10.6 Pre-Straining . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 45

10.7 Process Optimization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 45

10.8 In-Process Module Cleaning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47

10.8.1 Pump Off Cleaning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47

10.8.2 Forward Flushing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47

10.8.3 Reverse Flushing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47

10.9 Module Selection and Scale-up. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 48

11. Optional Equipment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50

11.1 Permeate Scale . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50

11.2 Backpressure Valve . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 54

12. Ordering Information . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 57

13. Contact Information - Worldwide . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 58


INTRODUCTION

1. IntroductionThe KrosFlo® Research IIi Tangential Flow Filtration (TFF) System is the ideal system for small-scale

process or R&D studies for microfiltration and ultrafiltration applications. The system consists of the

KrosFlo® Research IIi Pump and Pump Head, with an integrated KrosFlo® Digital Pressure Monitor,

and a disposable flow path, all of which have features that ensure efficient and reproducible TFF

processes. The Pump Drive used in conjunction with the KrosFlo® Research IIi Pump Head provides

flow rates from 0.01 to 2300 ml/min. The system has both audible and visual low and high pressure

alarms for the inlet and permeate, and a high pressure pump shut-off that help maintain membrane

integrity and achieve high product recovery. The KrosFlo Research IIi TFF system comes standard

with KF Comm, a software program that automatically downloads and graphs the run data including

inlet, retentate and transmembrane pressures and flow rates, into an MS Excel® spreadsheet. The

disposable flowpath including the filter, pressure transducers, tubing and fittings, eliminates the pos-

sibility of cross contamination and allows samples to be concentrated down to as low as 1 ml. Other

standard features include: a digital readout of the recirculation flow rate, the easy-to-use KrosFlo®

Research IIi Pump Head, and adjustable holders attached to the pump drive that secure the process

vessels, Spectrum’s HF modules and the entire flowpath.

The membrane surface areas range from 5 cm2 to 1050 cm2 and are available in 0.5 or 1.0 mm ID

Hollow Fibers made of Polysulfone (PS), Modified Polyethersulfone (mPES), or Mixed Cellulose Ester

(ME). Pore sizes range from 3 kD up to 0.5 µm for both Ultrafiltration and Microfiltration applications.

The KrosFlo® Research IIi TFF System offers many conveniences over traditional cross-flow mem-

brane systems. The hollow fiber (HF) membrane modules and cross-flow filtration system provides

faster and gentler separation that helps avoid membrane fouling and maximizes product recovery.

The disposability of the modules eliminates not only the potential for cross-contamination and costs

associated with cleaning and rinsing, but also the difficulties associated with validating reusable mem-

branes. Used in conjunction with HF membrane modules, the KrosFlo® Research IIi TFF System

offers the following advantages:

• Faster process times

• Superior filtration dynamics

• Module disposability

• Lower costs

• Direct and easy scale-up for production volumes

2. Safety Precautions

DANGER: High voltages exist and are accessible in the KrosFlo® Research IIi System.

Use extreme caution when servicing internal components.

WARNINGS: Tubing breakage may result in fluid being sprayed from pump. Use

appropriate measures to protect operator and equipment.

Turn drive off before removing or installing tubing. Fingers or loose clothing may get

caught in drive mechanism.

SAFETY PRECAUTIONS


CAUTION: Power must be turned off before connecting the external Octopus cable

to prevent damage to the drive.

CAUTION: To avoid electrical shock, the power cord protective grounding conductor

must be connected to ground. Not for operation in wet locations as defined by

EN 61010-1.

CAUTION: Do not contaminate the lubricant in the container, on the shaft or on the seal

with foreign material. Failure to observe this precaution may result in damage to the

seal and premature failure of the seal.

CAUTION: Risk of crushing. Keep fingers away from rotor while pump is in operation.

Stop pump before loading or unloading tubing.

Explanation of Symbols

CAUTION: Risk of Danger. Consult Operating Instructions for nature of hazard and

corrective actions.

CAUTION: Risk of electric shock. Consult Operating Instructions for nature of hazard and

corrective actions.

WARNING: PRODUCT USE LIMITATION

This product is not designed for, nor intended for use in patient connected applications; including, but

not limited to, medical and dental use, and accordingly has not been submitted for FDA approval.


SPECIFICATIONS

3. Specifications

OutputSpeed: 0.1 to 600 rpm

Torque output, Maximum: 180 oz-in (13 kg•cm)

540 oz-in Starting

Speed regulation: Line ±0.1% F.S.

Load ±0.1% F.S.

Drift ±0.1% F.S.

Display: 128 x 64 LCD w/ LED Backlight

Pressure Sensor Limits: -5.0 to 45.0 psi

InputSupply voltage limits: 90 to 260 Vrms @ 50/60 Hz

(Universal Input)

Current, max.: 2.2A @ 115 Vrms, or 1.1A @ 230 Vrms

ConstructionDimensions (L × W × H): 10.5 in × 8 in × 8 in

(267 × 203 × 203 mm)

Weight: 13 lb (5.9 kg)

Enclosure Rating: IP 33 per IEC 60529

EnvironmentTemperature, Operating: 0° to 40°C (32° to 104°F)

Temperature, Storage: –25° to 65°C (–13° to 149°F)

Humidity (non-condensing): 10% to 90%

Altitude: Less than 2000 m

Pollution Degree Pollution Degree 2

(Indoor use - lab, office)

Chemical Resistance: Exposed material is aluminum,

ABS plastic and vinyl

Pressure Sensors either

Polycarbonate (ACPM-499-03N) or

Polysulfone (ACPM-799-01N)

Compliance: (For ETL Mark);

UL 61010-1, CAN/CSA C22.2 No. 61010-1

(For CE Mark):

EN61010-1 (EU Low Voltage Directive) and

EN61326 (EU EMC Directive)

RoHS

SYSTEM CONFIGURATIONS AND MAJOR COMPONENTS


4. System Configurations and Major Components

4.1 KrosFlo® Research IIi Pump Drive, Pump Head & Integrated Pressure MonitorThe KrosFlo® Research IIi TFF System comes with the Digital Pressure Monitor integrated into the Pump

Drive as one unit. The Programmable Logic Controller (PLC) runs both of the Pump and Pressure mon-

itor functions. For further instructions on running the system, please refer to Section 10. The pump head

can accommodate tubing sizes from #13 (1/32” inner diameter ID) to #18 (3/8” ID) and can handle rates

from 0.01ml/min up to 2.3 loops per minute (LPM).

4.2 KrosFlo® Research IIi Mounting HardwareThe KrosFlo® Research IIi TFF System comes with an attached mounting plate for the KrosFlo®

Research IIi Pump Head and a stainless steel mounting bracket to connect the stainless steel hoop

post. The stainless steel hoop supports the module Trilobites (holders) and Reservoir Trilobites (hold-

ers) that come with the Starter Kit. The module Trilobites can accommodate Spectrum’s MicroKros®,

MidiKros®, MiniKros® Sampler/Sampler Plus, and some of the MiniKros® hollow fiber filtration mod-

ules. The reservoir Trilobites can secure reservoirs from 15 ml up to 2 L. A standard lab clamp is also

included to support larger reservoirs or filters as necessary.

4.3 KrosFlo® Research IIi Octopus Cable

The system comes with an ‘Octopus cable’ that has the following connections: 3 pressure trans-

ducers (feed, retentate, permeate), a USB port for capturing data on a computer, an RS232 port for

connecting a permeate Scale (optional) and a secondary RS232 port to connect the Automatic

Backpressure Valve (optional).

4.4 Flowpath KitThe Universal Flowpath Kit includes many different sized fittings for #13 tubing up to #17 tubing to

accommodate the three sizes of filtration modules used in the system: MicroKros® (1 ml – 50 ml),

MidiKros® (5 ml – 200 ml), and MiniKros® Sampler (50 ml – 4+ L). One each of 15 ml, 50 ml and 250

ml, 3-port conical bottom reservoirs are included. Packages of size #14 and #16 tubing are also

included. Filtration modules and other sized bottles or tubing sizes are available for purchase sepa-

rately. For specific instructions on flowpath construction and options, refer to Section 10.2,

“Assembling the Flowpath”.

4.5 Starter KitThe starter kit includes other items needed to construct and support the flowpath, including reusable

items such as: Reservoir and Module Trilobites, flow restrictors, tie wraps and a tie wrap gun.

5. System Options and Parts ListThe available universal system with corresponding catalog number is listed below:

KrosFlo® Research IIi TFF System Selection GuidePart Number Flowpath Type Pump Capacity TubingSYR2-U20-01N MicroKros, Midi, Sampler 2.3 LPM, 110/220V Pharmapure®, #14, #16

OPTIONAL EQUIPMENT:ACPC-F16-01N Automatic Backpressure Valve for use with #13,14 or 16 tubing

ACPC-F17-01N Automatic Backpressure Valve for use with #17 tubing

ACR2-SC4-01N Permeate Scale, 4 kg Capacity, 110V

ACR2-SC2-01N Permeate Scale, 4 kg Capacity, 220V

ACR2-SC5-01N Permeate Scale, 400g Capacity, 110V


SYSTEM OPTIONS AND PARTS LIST

ACR2-SC3-01N Permeate Scale, 400g Capacity, 220V

ACR2-SC8-01N Ohaus® Navigator Scale 10000g (1.0g), 110V




System Parts List

Part Description Quantity per System

KrosFlo® Research IIi Pump Drive w/ Integrated Pressure Monitor 1

Stainless Steel Hoop Posts w/ Mounting Screws 1

Attached Mounting Bracket for Pump Head 1

Octopus Cable 1

USB Extension Cable 1

Power supply Cable 1

Digital Pressure Transducers 3

KF COMM Data Collection Software Disc 1

KrosFlo® Research II Pump Head 1

Flowpath KitPlastic Fittings Box w/ Accessories 1

#14 Extended Life Pharmapure® Pump Tubing 25 ft

#16 Extended Life Pharmapure® Pump Tubing 25 ft

Starter KitTie Wrap Gun 1

Tie Wraps 100

Module Trilobite (holder) 2

Reservoir Trilobite (holder) 2

Pressure Control Valve

5/16” to 3/8” OD Tube 2

5/32” to 1/4” OD Tube 2

Reservoirs15 ml Conical Bottom with C-Flex® Molded Seal, 2 Diptubes plus Vent 1

50 ml Conical Bottom with C-Flex® Molded Seal, 2 Diptubes plus Vent 1

250 ml Conical Bottom with C-Flex® Molded Seal, 2 Diptubes plus Vent 1

6. Materials of Construction Part Description Material

KrosFlo® Research IIi TFF System (Product Contact Surfaces)

Tubing / Reservoir Closures C-Flex® / Pharmapure®

Reservoirs Polypropylene

Disposable Pressure Transducers Polysulfone

PUMP/PRESSURE MONITOR SET UP AND OPERATION


Plastic Fittings Polypropylene / Polycarbonate

Hollow Fiber Module

Membrane (1of 3) ME, mPES or PS

Housing Pigmented and Non-pigmented Polysulfone

Potting Polyurethane / Epoxy

End-caps Pigmented Polysulfone

7. KrosFlo Research IIi Pump/Pressure Monitor Set Up and Operation

7.1. Installation and Setup

Before Starting Drive

• The drive should be mounted on a flat horizontal surface, and no more than two (2)Pump Heads should be added for 600 rpm drives.

• The ambient air temperature should not exceed 104° F (40° C) and adequate air flowshould be provided for.

• The Octopus Cable should be attached to the back connector on the drive and thepressure sensors should be connected.

• Tubing should be clean and routed so that bend radii are at a minimum four (4) timesthe tube diameter and as short as possible.

CAUTION: Risk of crushing. Keep fingers away from rotor while pump is in operation.

Stop pump before loading or unloading tubing.

• Use a tube size of appropriate diameter for the flow rate and viscosity required.

• To maintain the best accuracy of flow rates, re-calibrate tubing regularly. See TubingCalibration in Chapter 7.2.4 Section C of this manual.

• For tubing selection and compatibility, see Tubing Selection Guide provided with thepump head.

• For Pump Head information, see Pump Head information in Chapter 8 of this manual.

• When cleaning or performing maintenance, please remove power from the drive.

DANGER: High voltages exist and are accessible in the KrosFlo® Research IIi System. Use

extreme caution when servicing internal components.


OPERATION

Mounting the Pump Head

Mount Pump Head and load tubing. Check to make sure that rollers are clean and free of defects.

7.2. Operation

7.2.1 Turning On the System

a. Plug the power cord into the IEC Connector, located on the rear of the drive. Plug the

opposite end of the power cord into an electrical outlet.

b. Flip the power switch located on the rear of the drive.

c. After the flash screen appears, the home screen will appear on the LCD screen.

CAUTION: To avoid electrical shock, the power cord protective grounding conductor must

be connected to ground. Not for operation in wet locations as defined by EN 61010-1.

WARNINGS: Tubing breakage may result in fluid being sprayed from pump. Use appropriate

measures to protect operator and equipment.

Turn drive off before removing or installing tubing. Fingers or loose clothing may get caught in drive

mechanism.

THE CONTROL PANEL


7.2.2. The Control Panel

• To navigate all menus on the drive, use the directional pad directly to the right of theLCD screen.

• The (ENTER) key, located in the middle of the directional pad, is used to enter a highlighted field or option. This key is often referred to as the ENTER key inthis manual.

• The (START/STOP) key, located at the top right of the control panel, is used tostart and pause the drive. This key is functional at any time, even when the display isin a setup screen. This key is often referred to as the START/STOP key in this manual.

• The ALARM OFF key, located at the bottom right of the control panel, is used tosilence the audible alarm.

7.2.3. Home Screen

A. Feed Pressure – Measurement from pressure sensor on the inlet or feed of the TFF filter.

Units can either be psi, mbar, or bar.

B. Permeate Pressure – Measurement from pressure sensor on the permeate of the TFF

filter. Units can either be psi, mbar, or bar.

C. Retentate Pressure – Measurement from pressure sensor on the outlet or retentate of the

TFF filter. Units can either be psi, mbar, or bar.

D. TMP (Transmembrane Pressure) – Calculated measurement of the average of the Feed

and Retenate pressures minus the Permeate pressure.


THE CONTROL PANEL

E. Feed Flow Rate – Recirculation flow rate based on calculation of pump rpm and select-

ed tubing size. Press the ENTER key when this is highlighted to change pump speed.

When the Feed Flow Rate screen is shown, the four pressures will still display on the right

side of the screen. Units are ml/min.

F. Setup – Pressing the ENTER key on this icon goes to the Setup screen. The Setup

screen has calibration functions, alarm settings, hardware setup and diagnostics. Please

refer to the Setup section below.

G. Pump Direction – Pressing the ENTER key on this icon toggles between clockwise and

counterclockwise flow direction.

H. Tubing Size – Pressing the ENTER key on this icon will select the tubing size (#13, #14,

#16, #25, HP15, #17, HP24, #18, HP35, #36, HP36).

I. Cursor Idle Place Holder – Place the cursor on this icon when cursor is idle to avoid any

unintended changes.

7.2.4. Setup Screen

A. Pressure CalibrateTo calibrate the pressure sensors, press the ENTER button when Pressure Calibrate is selected. Plug

a new pressure sensor (Spectrum P/N ACPM-499-03N or ACPM-799-01N) into each of the pressure

sensors to be calibrated. Use a calibrated pressure gauge in-line with the sensor to be calibrated and

attach a pressure source. Select the pressure for calibration and bring the pressure up to that point

with the pressure source. Select the sensor to be calibrated on the next screen (either Feed, Permeate

or Retentate). If FEED is selected, the Pressure Calibrate Feed screen will then appear:

Pf: The current reading for Feed Pressure

Set: Press the ENTER button to change the current reading to the new calibrated

value, as shown on the clibrated pressure gauge.

Clear: Press the ENTER button to clear all previous calibration values for the Feed sen-

sor. A confirmation screen will appear.

Cal: Press the ENTER button to confirm and save the new values.

Quit: Return to Setup Screen

B. Alarm Settings

THE CONTROL PANEL


To set the pressure alarms, press the ENTER button on the highlighted alarm. The pressure setting

for the alarm can then be changed using the arrows. Zero sets the alarm to off.

Pf Hi Stop: Secondary alarm that stops the pump, sounds a continuous beep and flashes the

screen when the Feed pressure reaches the setpoint

Pf Hi Alarm: Primary alarm that sounds a discontinuous beep and flashes the screen when

the Feed pressure reaches the setpoint

Pf Lo Alarm: Alarm that sounds a beep and flashes the screen when the Feed pressure

goes below setpoint

Pp Lo Alarm: Primary alarm that sounds a beep and flashes the screen when the Permeate

pressure goes below setpoint

Pp Lo Stop: Secondary Alarm that stops the pump, sounds a beep and flashes the screen

when the Permeate pressure goes below setpoint

Silent Alarm: Option to silence all audible alarms

C. Tubing Calibration

To Calibrate individual tubing sizes, follow these directions:

1. Mount Pump Head to drive

2. Insert appropriate tubing into Pump Head

3. Insert tube inlet into supply fluid

4. Insert tube outlet into desired container; container should be a graduated container or a

container placed on a scale for increased accuracy

If using a scale, an acceptable weight-to-volume conversion for water is 1 gram = 1 ml.


THE CONTROL PANEL

5. Turn on drive using power switch located on the rear of the drive.

6. Go to the Setup Menu by selecting the SETUP icon and pressing the ENTER key. Use

the UP and DOWN keys to highlight TUBING CALIBRATE in the Setup Menu and press

the ENTER key.

7. Set the drive for the desired flow direction, tube size, and flow rate. Note that these set-

tings are retained and transferred to other mode screens when entering or leaving the

TUBING CALIBRATION screen.

• The flow direction is set using the directional keypad to highlight the direction-al arrow. Pressing ENTER will toggle arrow between clockwise (CW) and coun-terclockwise (CCW).

• The tube size is set using the directional keypad to highlight the tube size field.Press ENTER and use the UP/DOWN keys to select the tube size. Press ENTERto SAVE the selection and return to TUBING CAL screen.

• The estimated flow rate is set using the directional keypad to highlight the flowrate field. Press ENTER and use the LEFT/RIGHT keys to select the digit to bechanged. Use the UP/DOWN keys to adjust the flow rate value. Select SAVE tosave settings, then select EXIT to finish. The drive will adjust this flow rate aftercalibration is complete.

• Note that the calibration volume is fixed and cannot be changed.

8. Press and hold the prime key on the drive console to prime the pump. Priming will stop

when key is released.

9. Place a measuring container at the pump outlet; highlight the START field and press the

ENTER key. The drive will run based on the default volume at the estimated flow rate

selected.

10. Upon completion of the calibration run period, the CAL VOLUME field will be highlight-

ed. Press the ENTER key and adjust the CAL VOLUME to the measured quantity. Use

the LEFT/RIGHT keys to select the digit to be changed, use the UP/DOWN keys to

adjust the value, and press ENTER to SAVE setting and EXIT the field.

A lower case “c” will display when the calibrated tubing size is selected. The volume units

are dependent on the flow rate units. For example, defining the flow rate unit as ml/min

will change the volume units to ml; defining the flow rate unit as oz/min will change the

volume units to oz.

11. To clear the calibration, select the CLEAR option on the tubing calibration screen when

the tubing to be cleared is selected.

Tubing Calibration Notes

• If the drive is stopped during calibration, empty the container and restart theprocedure.

• Calibration time at maximum allowable flow rate (default max flow rate) is 5-10seconds and at minimum allowable flow rate (approximately 4% of the maxi-mum flow rate) is 4 minutes.

THE CONTROL PANEL


• Minimum and maximum flow rates will change after a tubing calibration due toa re-calculation of the vol/rev.

• Optimum results are best obtained after tubing has been broken in by runningin pump for at least 10 minutes. Steps 8-10 can be repeated as necessary tooptimize the accuracy of the tubing cal.

CAL RUN TIME FORMULA

• 60 / (flow rate [ml/min] / cal volume [ml]) = cal run time (seconds)

INVALID CAL RUN TIME EXAMPLE

• tube size 13 flow rate range is 0.006 ml/min – 36.0 ml/min

• at flow rate of 1 ml/min, cal run time calculation is as follows:

60 / (1 ml/min / 6 ml) = 360 seconds

360 seconds exceed the max run time of 4 minutes (240 seconds)

D. Hardware Setup

Auto Tare: Select to turn on or off the auto tare function during system start up

Pressure Units: Select between psi, mbar, or bar

Time: Select to change system time

Date: Select to change system date

Key beep: Toggle key beep on/off

Display contrast: Raise or lower the screen contrast

Psi Decimals: Select number of decimals for psi units

Key Timeout: Select “Yes” or “No” to toggle on/off key timeout; when “Yes,” the key timeout will restore system to Home screen after 10 minutes of inactivity

Running Change: When on, the tubing size and pump direction can be changed while the

pump is running. If off, the pump needs to be stopped before the tubing

size or the direction can be changed.

Open Detect: The open detect function prevents the drive from turning when pump head

is in the open position. Pump head must be oriented vertically. In order to

use the pump head with the inlet/outlet to the left or right, the open detect

must be turned off.


TROUBLESHOOTING AND ERROR MESSAGES

Symptom Cause Remedy

Motor does not rotate, No Power 1. Check fuse; replace, if

Display does not light necessary

2. Check that unit is plugged

into a live line

3. Check connection of

power cord

4. Check the line cord for

continuity; replace if defective

5. Return for servicing

Motor Error #1: Motor OverCurrent

Description: Motor is drawing too much current for a shortduration of time

Error Condition(s): The motor current is above 4.0 A peak

Actions: Drive will stop immediately; determine if that pumphead is not binding and the load is above recom-mended maximum load; if error persists consultfactory

Motor Error #2: Mains Overvoltage

Description: The measured AC voltage reported by the drive is too high

Error Condition(s): The drive voltage is above 260V AC

Actions: Pump will stop immediately; check supply linevoltage, if error persists, consult factory

Motor Error #3: Mains UnderVoltage

Description: The measured AC voltage reported by the drive is too low

Error Condition(s): The drive voltage is below 90V AC

Actions: Pump will stop immediately; check supply linevoltage

NOTE: This error when is normal if displayed duringpower down; if error persists, consult factory

E. DiagnosticsFor factory use only.

7.2.5. Troubleshooting and Error Messages

TROUBLESHOOTING - ERROR MESSAGES


Motor Error #4: Motor OverTemp

Description: The motor is overheating

Error Condition(s): The temperature value from motor is above given threshold value

Actions: Pump will stop immediately; determine if ambient air temperature is less than 104° F (40° C), pump turns freely, and airflow is unrestricted; if error persists, consult factory

Motor Error #5: Communications Lost

Description: Communication to the motor is incorrect, has disappeared, or some other communi-cation error

Error Condition(s): Motor does not receive data from serial port

Actions: Drive will attempt to stop pump; power cycledrive; if error persists, consult factory

Checksum Error: ROM Checksum Wrong

Description: Run-time checksum (checked at power-on) contains a bad checksum value.

Error Condition(s): Checksum at power-on displays invalidvalue

Actions: Power cycle the drive; if error persists, con-sult factory

HW Error #1: Invalid Interrupt or Address

Description: Software jumps to an invalid address, invalidinterrupt, or other abort/exception (i.e., Data Fetch Abort, Software Prefetch Abort, Undefined Instruction, or FIQ Event)

Error Condition(s): May occur due to invalid pointer references, ram memory corruption, or other computer-related issues; the Abort Exception/Interrupt within the CPU is failing to branch out to its respective exception handler functions



TROUBLESHOOTING - ERROR MESSAGES

HW Error #2: Watchdog Timeout

Description: Program has stopped running (i.e., Software Locked Up) because Watchdog has not been updated

Error Condition(s): Interrupt triggered when the Watchdog has not been updated.



PUMP HEAD SET UP AND OPERATION

WARNING: PRODUCT USE LIMITATION

This product is not designed for, nor intended for use in patient connected applications; includ-

ing, but not limited to, medical and dental use, and accordingly has not been submitted for FDA

approval.

8. KrosFlo® Research IIi Pump Head Set Up and Operation

8.1 Safety Precautions

WARNING:

To reduce risk of electrical shock, connect only to a properly grounded receptacle.

Turn drive off before removing, adjusting or installing tubing. Fingers or loose clothing may

get caught in the rollers.

(The rotor is partially exposed when the Actuator lever is in the open position.)

Do not remove back cover. The strong spring inside may cause injury.

Tubing breakage may result in fluid being sprayed from pump. Use appropriate measures

to protect operator and equipment.

Risk of crushing. Keep fingers away from rotor while pump is in operation. Stop pump before

loading or unloading tubing.

Fingers may get trapped between rollers on the pump rotor.

Explanation of Symbols

WARNING: This word and symbol are needed in safety messages for hazards that are not immediately

accessible, but that present a probability of serious personal injury if the hazards are not

avoided. These safety messages describe precautions that must be taken to avoid hazards.

WARNING Warning or Danger with this symbol indicates probability of serious injury exists if crushing

hazard is not avoided.or DANGER:

PUMP HEAD SPECIFICATIONS


Number of rollers: 3

Maximum pump speed (rpm): 600

Maximum torque load—Starting: 400 oz-in (29 kg-cm)

Maximum torque load—Running: Up to 180 oz-in (13 kg-cm)

Housing materials: Glass-filled polypropylene (PP), polyphenylene

sulfide (PPS), nylon (PA)

Roller materials: Stainless steel (SS)

Bearing materials: Sealed stainless steel

Rotor materials: Stainless steel

Chemical resistance: Most substances, except strong acids or alkalis,

organic solvents or hydrocarbons.

Temperature, Operating ‡: 32°F to 104°F (0°C to 40°C)

Temperature, Storage : –49°F to 149°F (–45°C to 65°C)

Humidity: 10% to 90% (non-condensing)

Altitude: 2000 m or less

Dimensions (W x H x D): (Excluding shaft and cosmetic cover)

Operating 3.45" x 4.75" x 3.08" (8.8 cm x 12.1 cm x 7.8 cm)

Open 3.45" x 5.65" x 3.08" (8.8 cm x 14.4 cm x 7.8 cm)

Weight: 1.1 lb (0.5 kg)

*For NORPRENE®, PHARMED®, and TYGON® tubing. Values will be less with silicone, C-FLEX®, and Viton-®. Flow rate and discharge

pressure will vary based on tubing size, formulation, and operating temperature. The chart above is only a guide.

‡ Use in this temperature range for continuous duty operation with no decrease in performance or product life. Pump heads will work

outside this range with some possible reduction in performance or product life.

8.2 Specifications

Typical Flow, Pressure and Vacuum Data—3 roller pumps

LS® Flow rate* Discharge Pressure* Vacuum* Suction Lift*

Tubing @ 1 rpm @ 600 rpm Continuous Intermittent @ 600 rpm @ 600 rpmmL/Rev mL/min psig (bar) psig (bar) in (mm) Hg ft (m) H20

L/S®

13 0.06 36 25 (1.7) 40 (2.7) 26 (660) 29 (8.8)

L/S®

14 0.22 130 25 (1.7) 40 (2.7) 26 (660) 29 (8.8)

L/S®

16 0.80 480 25 (1.7) 40 (2.7) 26 (660) 29 (8.8)

L/S®

25 1.7 1000 20 (1.4) 35 (2.4) 26 (660) 29 (8.8)

L/S®

17 2.8 1700 15 (1.0) 20 (1.4) 20 (510) 22 (6.7)

L/S®

18 3.8 2300 10 (0.7) 15 (1.0) 20 (510) 22 (6.7)

Part Number Roller and Bearing Material Number of Rollers MasterFlex® L/S® Tubing

ACR2-H3I-01N Stainless Steel 3 L/S®

13, 14, 16, 25, 17, 18

MasterFlex®L/S®

Tubing


PUMP HEAD INTRODUCTION

8.3 IntroductionThe KrosFlo® Research IIi Pump Head (part number ACR2-H3S-01N) provides a simple, easy-to-use peristaltic pump system whencombined with a KrosFlo® Research IIi Pump Drive. The pump headaccepts several different tubing sizes for a wide range of flow rates.The unique lever actuator design and automatic tubing retentionallow for quick tubing changes. The Pump Drive is designed totransfer or dispense liquids of various viscosities, as well as solidssuspended in liquids. It is NOT intended to pump flammable orexplosive substances.

A mounting kit adapts this pump head to the Pump Drive. The kitincludes a mounting plate and four screws, which are used to attachthe mounting plate to the drive. The pump head is quickly and easi-ly attached to the mounting plate or another pump head by meansof the bayonet feature on the back of the pump head. The pumphead may be oriented with tubing entering and exiting to the left,right, up or down. In order to mount the pump head in the left, right,or up orientations, the Open Detect function must be turned off. Seethe Hardware Setup section under the Setup menu. Mounting orien-tation options may be limited with some drives. Pump heads are eas-ily removed using the one-finger release lever (Bayonet Lock Lever)on the upper left side of the pump head (see Figure 2). The tang onthe pump head shaft couples the pump head to standard pump dri-ves to transmit power.

Each KrosFlo® Research IIi Pump Drive is supplied with one mount-ing kit pre-assembled.

Figure 4 Bayonet feature locked in horizontal pump orientation

Approximately 30°

Figure 1 Attaching mounting plate to drive

(4) Screws

Bayonet Tabs Mounting Plate

Figure 2 Back of Research II Pump Head

Pump Shaft with Tang

BayonetFeatures

Bayonet LockLever, locked

from rearposition

Figure 3 Position for engaging bayonet feature for horizontal mounting

8.4 Installation and Removal

1. If mounting plate is not attached to the pump drive, attach it using

the provided four Phillips head screws (see Figure 1).

2. Orient the pump head with its back facing the drive and insert the

tang on the pump head shaft into the shaft’s slot on the drive. Align

the bayonet features on the back of the pump head with the bayo-

net tabs on the front of the mounting plate (see Figures 1, 2 and 3).

The pump head should be tilted about 30°counterclockwise from

the intended installed orientation. Press pump head firmly against

the drive and rotate clockwise until no more rotation is possible (see

Figure 4). The bayonet lock lever will automatically snap toward the

back of the pump, locking it to the mounting plate.

3. Remove the pump head from the drive by holding the bayonet lock

lever forward while rotating the pump head as far as possible in the

counterclockwise direction, then pull the pump head away from the

drive to detach it. The actuator lever should be in the far right posi-

tion when removing the pump head (see Figure 5).

WARNING: Stop the pump drive before installing or removing the pump head from the drive.

PUMP HEAD TUBING LOADING AND UNLOADING


8.5 Tubing Loading and Unloading

1. To load tubing, open the pump head by moving the actuator

lever counterclockwise (see Figures 5 and 6). Insert a loop of

tubing into one open tubing retainer, between the occlusion

bed and the rollers and into the other tubing retainer (see

Figure 7). Position the tubing so that it is firmly centered

against the rollers. While holding the tubing ends, move the

actuator lever back to the far clockwise (right) position, as

shown in Figure 5. The pump head will automatically grip the

tubing. Tubing sizes L/S® 13, L/S® 14, L/S® 16, L/S® 25,

L/S® 17 and L/S® 18 (thin wall) will automatically be

stretched by the pump. Approximately 5 pounds of force

must be applied to the actuator lever to fully close the pump

head and place the lever in its locked position (far right posi-

tion) or to fully open the pump head (far left position).

NOTE: It is unnecessary to have an end of the tubing free to load or

unload tubing from the pump head. A length of tubing may be

loaded into the pump without disconnecting it from adjacent

devices.

WARNING: Stop the pump drive before installing or removing tubing from the pump head.

Figure 6 Pump head in fully open position

Actuator Lever-Far Left Position Occlusion Bed

Roller

Figure 5 Pump head in fully closed position

Bayonet Lock Lever-PullForward

to Release

Actuator Lever-Far

Right Position

Figure 7 Tubing path through pump head - during loading

Tubing

Retainer

(one per side)

2. Before unloading tubing

from the pump head, first

turn off the drive. Open

the pump head by mov-

ing the actuator lever

counterclockwise (left),

as described above. This

will automatically open

the tubing retainers, as

well as lift the occlusion

bed away from the tub-

ing. Pull the tubing away

from the pump head.

Occlusion

Bed

NOTE: When pump is not being

used, store with actuator

lever half way between far

left and far right positions

(see Figures 5 and 6).


PUMP HEAD MULTI-CHANNEL MOUNTING

8.6 Multi-Channel Mounting1. The KrosFlo® Research IIi Pump Heads can be mounted in

tandem (see Figures 8, 9 and 10). Once the mounting plate

is attached to the pump drive, no other mounting hardware

is required.

2. Install the first pump head according to the mounting

instructions in Step 4.2

3. To install a second pump head, the cosmetic cover must be

removed from the first pump head. Grasp the cover by the

notches and pull it off (see Figure 8). Align the second pump

head to the first, as if the first pump head were the drive, and

continue to follow pump head mounting instructions in step

4.2 (see Figures 9 and 10).

NOTE: The tubing on the

inner pump head(s)

can be changed

without removing

the outer pump

head(s) from the

drive.

Figure 8 Preparation to mount a second

pump head

Cosmetic Cover

Notch (here and on opposite edge)

Bayonet Tabs on Pump Front

Figure 9 Engaging bayonet of second pump head to bayonet tabs on first pump head

CAUTION: Be sure that bayonet features on back of each pump head are fully engaged with

bayonet tabs on the mounting plate or adjacent pump head before operating

pump drive. Bayonet lock lever (see Figure 5.) will snap back when bayonet

features engage completely.

Figure 10 Drive with both pump heads locked in position

8.7 Maintenance and Cleaning

No lubrication is required for the KrosFlo® Research IIi Pump Head. Only use a mild detergent solution or

70% isopropyl alcohol to clean the pump head. Do not immerse nor use excessive fluid. Pump head

requires no maintenance beyond cleaning. There are no user serviceable or replaceable parts inside.


KF COMM SOFTWARE

9. KF Comm Software

9.1 KF Comm License

Copyright © 2010 Spectrum Laboratories, Inc., Rancho Dominguez, USA. All rights reserved.

Redistribution and use in source and binary forms, with or without modification, are permitted provided that

the following conditions are met:

• Redistribution of source code must retain the above copyright notice, this list of conditions and the following disclaimer.

• Redistribution in binary form must reproduce the above copyright notice, this list of conditions and the following disclaimer in the documentation and/or other materials provided with the distribution.

• Neither the name of Spectrum Laboratories, Inc. nor the names of its contributors may be used to endorse or promote products derived from this software without specific prior written permission.

THIS SOFTWARE IS PROVIDED BY THE COPYRIGHT HOLDERS AND CONTRIBUTORS “AS IS” AND ANY

EXPRESS OR IMPLIED WARRANTIES, INCLUDING, BUT NOT LIMITED TO, THE IMPLIED WARRANTIES OF

MERCHANTABILITY AND FITNESS FOR A PARTICULAR PURPOSE ARE DISCLAIMED. IN NO EVENT SHALL

THE COPYRIGHT OWNER OR CONTRIBUTORS BE LIABLE FOR ANY DIRECT, INDIRECT, INCIDENTAL, SPE-

CIAL, EXEMPLARY, OR CONSEQUENTIAL DAMAGES (INCLUDING, BUT NOT LIMITED TO, PROCUREMENT

OF SUBSTITUTE GOODS OR SERVICES; LOSS OF USE, DATA, OR PROFITS; OR BUSINESS INTERRUPTION)

HOWEVER CAUSED AND ON ANY THEORY OF LIABILITY, WHETHER IN CONTRACT, STRICT LIABILITY, OR

TORT (INCLUDING NEGLIGENCE OR OTHERWISE) ARISING IN ANY WAY OUT OF THE USE OF THIS SOFT-

WARE, EVEN IF ADVISED OF THE POSSIBILITY OF SUCH DAMAGE.

KF COMM SOFTWARE - INSTALLATION


9.2. Installation

9.2.1 KF COMM Software InstallationThe KrosFlo® Research IIi TFF System is accompanied with a disc of software that can be used on Windows®

PCs. The software requires Microsoft® Windows 95 or newer and Microsoft® Excel® 32-bit 2003 or newer.

KF COMM will work properly on a 64-bit computer if Excel® 32-bit is used to run the software.

The supplied software includes an ActiveX control which allows other applications to communicate with the

Pressure Monitor, a spreadsheet template for automatically recording your process separation conditions, and

a spreadsheet for assigning COM ports. The Pressure Monitor and KrosFlo® Research IIi TFF System acces-

sories are connected to the computer with a USB connection. The USB connection creates a virtual serial

(COM) port that the software connects to.

While there are ways to share serial ports across computers, Spectrum has not tested doing so. The spread-

sheet templates installed with KF COMM may be copied and moved to other locations on the Windows® PC,

and will continue to retain its KF COMM capabilities. However, if the spreadsheets are moved to a Windows®

PC without KF COMM installed, the spreadsheets will not have KF COMM capabilities. However, it can be

viewed and manipulated like a typical Excel® sheet on computers without KF COMM.

To start the software installation, put the software disc into the CD drive and it will load automatically. If it

does not start, double-click on the icon for the CD drive and then double click on the Setup.exe item (the

Setup.exe item looks like ).

After the installation begins, accept the terms of the software license to continue. The license terms

are listed in Section 9.1 of this manual.

You will be prompted to select the instllation location (typically C:/Program Files/Spectrum

Laboratories). One copy of each spreadsheet will be installed in this location. The KF Comm control

will be installed in your windows system directory (typically C:\Windows\System).

If the user does not have administrator privileges on the computer, the installation will fail without

giving notice that it has not completed successfully.

9.2.2 Drivers for USB connectionTo use the USB connector, attach the USB extension cable from the Octopus cable wire labeled

“Computer” to a PC. Windows® will automatically search for the proper driver if the system is turned on. If

not found, a prompt will be displayed to find the driver location. The driver is located in the CD that comes

with the system (folder: FTDI USB Drivers). The FTDI driver can also be downloaded and installed from

http://www.ftdichip.com/Drivers/VCP.htm (the device is FT2232). Windows® will then perform a similar

search for the driver for the permeate scale even if the scale is not attached or not going to be used.

9.3. Instructions for Use

9.3.1 Opening the Template

The workbook template used to record the process parameters is named TFF Trial Template. This

Excel® workbook contains programming that automatically collects and graphs real-time run data.

Other files located in the folder include: the KrosFlo Research IIi System Manual, an Excel® file

called Override and another Excel® file called KMP System. The Override file assigns the COM port

for the USB data connection from the KrosFlo Research IIi System to the PC. This is done if the

COM ports cannot be detected from the TFF Trial Template spreadsheet. The KMP System is used

for data collection with Spectrum Labs’ larger KrosFlo® MiniKros Pilot TFF System.


KF COMM SOFTWARE - INSTRUCTIONS FOR USE

The preset macros record pressure and flow readings during the experiment. Depending on the

security settings and the version of Excel®, a prompt may ask for permission to run the macros

upon opening the template. If the Excel® security level is High, check the “Always trust macros from

this source” box before clicking the “Enable Macros” button. It may be necessary to have an

administrator accept the certificate used to sign the macros.

If the macros are not enabled, the data will not be collected. If opening a workbook that already

contains data, the data remains accessible whether or not macros are enabled.

9.3.2 Data Collection Tools The TFF Trial Template contains both data collection worksheets and automatic graph worksheets (see

Section 9.3.3 below). When one of the data collection worksheets is open and the KrosFlo Research IIi

System is connected to the computer, the first row of the spreadsheet will display the current readings (the

yellow background cells next to the “Now”). If the system is connected and on, yet the first line remains

blank, use the configure control to find the COM port.

The first four icons on the standard toolbar can be used to control the data collection. The same items will

also appear on the Data menu at the top of the window. For newer versions of Excel, the icons appear in

the Add-Ins menu.

Configure Collection: Used to set how the data is collected. Clicking on this will bring up a dia-

log. Use this dialog to set the number of seconds per line collected (default is 60 sec-

onds per line). A 3 hour run at 1 line per second would produce about 10,000 data

points. To collect fewer points, a larger number would need to be set for the seconds per

line parameter. An Excel sheet can only contain about 65,000 rows; this limits collection

to about 18 hours at 1 second per line.

This control also can be used to find the COM port for the USB connection from the

Octopus Cable. If the COM port is unknown, click the Find Comm button. The control

will cycle through the free COM ports on the computer to identify the KrosFlo Research

IIi System. If the system is turned on and connected to the computer through the USB

or Serial connection (from the Valve serial connector), the COM port should be found

within 1 cycle of the available ports. The control will continuously seek the Pressure

Monitor until one is found or until you click the Cancel button. If another program is being

used to communicate with the system, you'll need to close that program before select-

ing the Configure tool so that the serial port becomes available to the workbook.

Start Collection: Used to start collecting data. Once the first row in the worksheet is properly dis-

playing the real-time pressure information, click on this icon to begin saving pressure

data. After clicking on the Start Collection button, the Configure Collection will be dis-



abled until Stop Collection is pressed.

Stop Collection: Used to temporarily stop collecting the pressure data. After clicking on Start

Collection to start saving data, the Stop Collection is used to suspend data collection. If

you select Start, Stop, and then Start again, collection will resume where it left off; it does

not begin again at the top of the sheet.

End Collection: Used to end data collection. Certain functions in Excel® need to be disabled for

the data in the TFF Trial Template to be accessible. Clicking this will stop the spreadsheet

from updating the first line, stop the data collection, and disable the other controls. The

data can then be accessed for analysis.

If the End Collection button is clicked once and then the workbook is saved, data col-

lection will still be available the next time the workbook is opened. If the End Collection

button is clicked twice and then the workbook is saved, data collection will no longer be

available the next time the workbook is opened.

9.3.3 Navigating the Worksheets and Charts

1. Worksheets Overview

In the TFF Trial Template workbook, there are three different worksheets for data collec-

tion (orange tabs at bottom of Excel® workbook). These three worksheets are used for

collecting data for different stages of a filtration process:

Integrity Test – For performing test to ensure membrane integrity

Module Characteristics – For analyzing initial water flux

Trial Data – For collecting run data

NOTE: Data is only collected in the worksheet that is open.

The six remaining worksheets with the green tabs are automatic graphs. Here is a brief overview:

Graph Description

NWP Graphs data from ‘Module Characteristics’

(Normalized Water Permeability) worksheet for analyzing initial water flux

CF and Pressures vs. VT

Flux Rate vs. Time

Flux vs. CF Graphs data from ‘Trial Data’ worksheet

Feed Flow Rate vs. Time to analyze process conditions

Pressures vs. Time

!!!!!



2. Header Information and Options

The data collection worksheets have a header (see above) with various inputs for filter selection, run

options, and run data — including:

a. Date

b. Filter Module part number

• When the filter part number is entered, the filter information — including the surface

area, approximate fiber count, fiber length, shear rate guidelines and membrane type —

will be automatically entered.

c. Serial number

d. Tubing size through the pump head and Tubing Calibration

• Correct tubing size must be selected for proper feed flow rates to be captured

in the spreadsheet.

• If the pump is calibrated (see 7.2.4 section C), then the correction factor will

need to be adjusted to determine the correct feed rate. This is done while the pump

is running. Click on the icon next to the Pump Calibration value and enter the value

in the displayed flow rate on the pump display.

e. The starting amount of material for Concentration Factor calculation

f. The method for permeate collection – by weight or by volume

• When using the optional permeate scale for automatic data collection, the

“By Weight” button must be selected.

g. Filtration application, objective and notes

h. PSI vs. mbar units for pressure

i. Shear rate guidelines

• Feed rate @ 4000s-1 is the recommended maximum feed flow for shear sensitive

applications. Feed Rate @ 12000s-1 is the recommended maximum feed flow for

fouling applications.

j. Average Permeate Flux and Permeate Flow over the entire filtration run

k. Module Table Date

• Date reference for when the filter module database created. Database will be

updated as new filters and filter membranes are released.

!!!!



3. Module Characteristics WorksheetThe ‘Module Characteristics’ worksheet collects data to measure the NWP of the filter.

The header for the data collection in the worksheet looks like this:

Here is a breakdown of the abbreviation descriptions:

Column Abbreviation Description

Time Time (H:MM:SS)

Pinlet Inlet Pressure

Pretentate Retentate Pressure

Ppermeate Permeate Pressure

TMP Calculated Transmembrane Pressure

DP Pressure Drop through filter

Qinlet Feed Flow (calculated from Pump RPM and tubing size)

Qpermeate Permeate Flow (measured and input by user at specific times)

Temp Temperature (measured and input by user at specific times)

TCF Temperature Correction Factor (calculated from temperature)

Water Flux Liters of permeate per m2 of surface area per hour

(calculated from permeate flow/SA)

Water Flux @ 20°C Corrected Water Flux

WP Water Permeability

NWP Normalized Water Permeability

NOTE: For the data collection worksheets, there are three different types of columns: collected data (yellow), calculated data (light green) and user input data (tan).

After a filter has been properly rinsed and/or wetted according to the instructions on the

insert titled, “Hollow Fiber Filter Preparation Guide,” the initial water flux data can be mea-

sured. After making sure that the first row is reading the current pressure data, click on

the Start Collection button. At the set time interval, make a permeate flux measurement

by using a graduated cylinder and input the data in the Qpermeate column. Continue col-

lecting data points at different transmembrane pressures (TMP’s). Input the temperature.

The ‘NWP’ worksheet will automatically graph the NWP vs. TMP.

!!!!



4. Integrity Test Worksheet

The ‘Integrity Test’ worksheet measures the inlet pressure at set time intervals and cal-

culates the ∆P over time. If the psi/min change is not > 0.5 psi/min then the filter is fit for

use. A sharp decrease in pressure over time indicates either a broken filter or incom-

pletely wetted filter. Follow the instructions in the “Hollow Fiber Filter Preparation” insert

and/or Section 10.3 for proper wetting and integrity testing procedures.

5. Trial Data Worksheet

The Trial Data worksheet is used to log and calculate the filtration run data (see sample

below).

The columns are the same as the ‘Module Characteristics’ with the following additions:

Column Abbreviation Description

Qretentate Retentate Flow

Mpermeate Total mass of permeate (user inputted). This can be changed to total

volume of permeate by clicking on the ‘By Volume’ button on the header.

LMH Filtrate rate in liters per m2 of membrane per hour

VT Volumetric Throughput

(a measure of the total permeate per cm2 of SA)

Pump RPM of the pump

Temp Temperature (user entered)

Conc. Factor Concentration Factor

Shear Shear rate of the fluid through the fibers (s-1)

NOTE: The permeate volume/mass are to be measured and inputted by the user at the corre-

sponding times. The spreadsheet will then calculate the permeate flow rate, the LMH and

the VT. If using the Permeate Scale accessory, the data will be automatically entered into

the worksheet and all the calculations will be completed. Please refer to the Permeate

Scale section of this manual for the installation and use of the scale (see Section 11.1).

9.3.4 CompliancySpectrum offers an additional software program that enables the KF COMM program to comply with 21

CFR Part 11 regulations. Please contact Spectrum at 1-(800)634–3300 for further information.

!!!!!!!!


BASIC CONCEPTS OF TANGENTIAL FLOW FILTRATION

10. Basic Concepts of Tangential Flow Filtration (TFF)Membranes use the principle of barrier separations to differentiate components based on size.Components larger than the membrane pore are quantitatively held back by the membrane whilesmaller components pass through the membrane structure along with the permeate. Although thereare other methods for driving the separation process — such as electric charge (e.g. caustic-chlorinecells) and diffusion (e.g. dialysis and oxygenation devices) — Spectrum hollow fiber membrane mod-ules are designed for pressure-driven applications.

10.1.1 Dead-end FiltrationTraditional sieve (or dead-end) filtrations consist of forcing a solution containing suspended solidsdirectly through the membrane structure. Solids retained by the membrane collect on the surface ofthe membrane media, continually reducing the permeation rate and eventually plugging the device.See illustration below.

The driving force for permeation is the pressure difference between the feed and the permeate, calledthe transmembrane pressure (TMP). For dead end filtration, this is given by the following equation:

PTMP = Pfeed – Ppermeate

10.1.2 Tangential Flow FiltrationTangential Flow Filtration (TFF) is an efficient way to separate streams that would quickly becomeplugged if processed by dead-end techniques. When using tangential flow techniques, most of theprocess fluid flows along the membrane surface rather than passing through the membrane structure.Fluid is pumped at a relatively high velocity parallel to the membrane surface. Except for water treat-ment applications, only a small percentage of the tangential flow along the membrane surface endsup as permeate. In most cell and particle separations, only 1 – 5% of the inlet flow to the membranedevice becomes permeate. The remaining 95 – 99% exits the membrane device as “retentate”. Theretentate is recirculated back to the process reservoir and the module inlet such that another 1% to5% can be removed as permeate. This recirculation process continues in rapid succession, generat-ing a significant and continuous permeation rate. See the illustration below.

BASIC CONCEPTS OF TANGENTIAL FLOW FILTRATION


Some examples of tangential flow membrane geometries include: stacked plate and spiral devicesthat utilize flat sheet membranes, tubular devices, along with shell and tube devices that use hollowfiber membranes. In the case of tangential flow separations, the driving force is the TMP, the differ-ence between the average of the module feed and retentate pressures and the permeate pressure:

PTMP = (Pfeed + Pretentate) / 2 – Ppermeate

Filtrate flow results in a build up of retained components on the membrane inner lumen surface.Generally, these components are carried down the length of the hollow fiber and out the end of themodule by the sweeping action of the recirculating fluid. Occasionally, a cake layer may accumulateon the surface of the membrane. This boundary layer is composed of solids and/or solute macro-molecules which are retained by the membrane from filtration. This phenomenon, often erroneouslyreferred to as “concentration polarization,” can affect module performance by reducing the function-al size of the membrane pore. In other words, the cake layer becomes the membrane barrier, a“dynamic membrane.”

Caking is influenced by fluid variables: degree of solvation, concentration and nature of the solids andsolutes, fluid temperature, along with operating cariables — such as solution velocity along the mem-brane — and TMP (refer to Section 10.8). Controlling this phenomenon is the key to maximizing flux,solute passage, and optimizing the process parameters. Caking can usually be controlled by ensuringadequate fluid velocity at the liquid-membrane wall interface. Fluid velocity is controlled by the pump-ing rate. Recirculation rates depend on the quantity of fibers in a module and shear rate considera-tions. Typically, a shear rate of 12,000 s-1 is used for filtration applications and up to 4,000 s-1 is usedfor perfusion applications. These rates are guidelines only and should be optimized for each processapplication. Certain applications may work well at reduced rates while others may require rates that aresignificantly higher. When protein passage through the membrane structure is important, particularattention should be paid to feed or recirculation rate. In general, high feed rates allow more efficientprotein passage. Depending on the characteristics of the retained components (cells, cell debris, diag-nostic particles, etc.), a caking layer can form on the membrane wall that is actually tighter than themembrane pores. In these instances, high recirculation rate and low transmembrane pressure oftenhelp. Variables such as viscosity or shear sensitivity of the solution components may prevent the userfrom generating enough velocity to reduce membrane caking. Often in these cases, applying a deter-mined amount of back pressure on the permeate helps to prevent caking. Cell perfusion applicationsfrequently use permeate back pressure to prevent caking.


ASSEMBLING THE FLOWPATH

10.2 Assembling the Flowpath

The following are the instructions for assembling the KrosFlo® Research IIiTFF System flowpaths.Figure 11 is given as a representative layout of the flowpath. For a description of the differencebetween Constant Volume Mode and Batch Mode, refer to Section 10.4.

10.2.1 Constant Volume ModeRefer to Figures 12, 13 and 14 for assembling either one of the three different size flowpaths(MiniKros® Sampler, MidiKros® and MicroKros®, respectively) for constant volume mode.

1. Attach one Reservoir Trilobite (holder) to the bottom end of the right post on either sideof the KrosFlo® Research IIi Pump Drive. The Reservoir Trilobites should be oriented suchthat the front (wide) groove faces forward and its respective thumb screw is directedaway from the pump. Attach both Module Trilobites to the left post above the ReservoirTrilobite. Reservoir and Module Trilobites can be attached to the posts by hooking theposts into the groove on the back (narrow) end of the Trilobite and securing the Trilobitesin place with the back thumb screw (Figures 11A and 11B).

2. Attach the HF filtration module by inserting it into the front groove on both the ModuleTrilobites. Slide the Trilobites away from each other until the module is held firmly in place.Secure the Trilobites by tightening the back thumb screws (Figure 11B).

3. Attach the smaller processing reservoir by inserting it into the front groove of theReservoir Trilobite on the right or left side of the pump. Secure in place by tightening thefront corresponding thumb screw (Figure 11B).

4. A secondary Reservoir Trilobite or standard lab clamp can be used to secure the bufferreservoir (Figure 11B).

NOTE: The reservoir Trilobites are designed with a thumb screw to hold the smaller reservoirs

by either the bottle or cap, and to hold the large reservoirs by the cap only. The smaller

reservoirs can be held suspended above the surface of the bench top, while the larger

reservoirs need to rest on the bench top.

5. Assemble the flowpath as shown in Figures 12, 13 and 14 for the MiniKros Sampler (orMiniKros), MidiKros and the MicroKros, respectively. The Universal Flowpath kit comeswith 3 different sizes of plastic fittings to make the necessary connections. These include1⁄16” Hose Barb fittings for tubing sizes 13 and 14 used for the MicroKros or MidiKros, 1⁄8”Hose Barb fittings for tubing size 16 used for MidiKros and 1⁄4” Hose Barb fittings for tub-ing size 17 used for the MiniKros Sampler or MiniKros filters.

The reservoirs that come with the system include the 15 ml, 50 ml and 250 ml. All havethree ports. The middle port is used as the inlet (flowing into the filter), the other diptubeis used as the retentate (returning from the filter), and the third port without any diptubeis used as the vent/buffer addition line. The 15 ml reservoir is typically used with #13 tub-ing and MicroKros filters for small volume applications (1-50 mls). The 50 ml reservoir istypically used with #14 tubing and MidiKros modules for 5-1000 mls. The 250 ml reser-voir is typically used with #16 tubing or #17 tubing using the 1⁄8” to ¼” hose barb adap-tors for MiniKros filters (30 ml -10 L). A 500 ml reservoir (ACBT-500-C1N) is availablewith ¼” diptubes if larger flow rates are needed.

6. Connect the appropriate size HB x HB connector to the tubing coming out of the bufferreservoir (#6 in Figures 12, 13 and 14) and connect it to the vent of the process reser-voir (#5 in Figures 12, 13 and 14).

OVERALL FLOWPATH: FRONT AND TOP VIEW


10001000

MODULETRILOBITE(HOLDER)

RESERVOIRTRILOBITE(HOLDER)

Figure 11A. System Schematic - Top View

Figure 11B. Overall Flowpath - Front View


NOTE: The buffer line can also be connected to a Tee in the retentate line (#4) for adding inthe wash buffer. This can be done to increase mixing efficiency. During the filtration,make sure the vent is closed and open the permeate line. Ensure that a vacuum hasbeen created in the closed system before opening the pinch clamp to the buffer reser-voir. The inlet pressure will decrease if the vacuum is created. If a vacuum is not creat-ed, the sample has the potential of flowing into the buffer reservoir instead of the recir-culation reservoir.

7. Assemble the rest of the flowpath — including pressure transducers and HF filtrationmodule for MiniKros® Sampler, MidiKros® and MicroKros® — according to Figures 12,13, and 14 respectively, and the following steps:

MiniKros® Sampler Flowpath (Figure 12, page 35)

i. Cut an appropriate length of #17, ¼’’ ID tubing (ACTU-P17-25N, not included with system) to run from the process reservoir (#5 Figure 12) through the pump head (#7) andto the inlet of the filter. Attach one end of tubing to a ¼’’ HB x ¼’’ HB connector andattach the connector to the tubing on the 500 ml process reservoir (ACBT-500-C1N)that corresponds to the lowest diptube.

NOTE: The 250 ml process reservoir can be used in place of the 500 ml if the 1⁄8’’ HB x 1⁄4’’

HB connector is used to connect the diptubes to the inlet #17 tubing.

Place tubing through the pump head and then connect other end of tubing to the Pro-Connex fitting (HB x TC x female Luer™). For filters with hose barb fittings, use the ¼”HB x ¼’’ HB x Luer pressure transducer tee connector (not shown).

ii. Cut another length of tubing (#4) for the outlet of the filter through the AutomaticBackpressure valve (if used) to the other ¼’’ diptube on the 500 ml process reservoir.

iii. Close off the lower permeate side port of the HF module with either a male Luer cap(provided with module), or a short piece of tubing and a HB plug.

iv. Connect a short piece of tubing from the upper HB permeate side port (use Male Luerx HB fitting provided with module if necessary) to a pressure transducer connector (¼”HB x ¼” HB x Female luer) for the permeate pressure transducer. Connect an appropriate length of tubing from the HB-free end of the tee connector to the desiredpermeate collection vessel (not provided). Slide a pinch clamp (#9) over the permeatetubing.

v. Attach pressure transducers (#1) to the inlet, retentate and permeate pressure Pro-Connex or transducer tee connectors. Plug the pressure transducer cables intothe appropriate labeled jacks on the Octopus cable at the rear of the pump.

vi. Attach a buffer reservoir (#6) to the buffer addition port (1⁄8” ID tubing) on the 500 mlreservoir for constant volume diafiltration mode.

vii. Attach a vent line stopcock or pinch clamp to open/close vent.

viii. If preferred, suspended tubing can be secured in the tubing notches on the sides ofthe module and reservoir Trilobites.

ix. All HB connections may be secured with tie wraps and a tie wrap gun.


MINIKROS® SAMPLER FLOWPATH


Figure 12. MiniKros® Sampler Flowpath

LEGEND:1. Polysulfone Pressure Sensor (ACPM-799-01N)2. Pro-Connex® TC x HB x Luer Pressure sensor connector 3. Manual Backpressure clamp4. Retentate Line (#16 or #17 tubing)5. 500 ml 4 Port Process Reservoir (ACBT-500-C1N) 6. Buffer Reservoir7. Feed Pump8. Permeate Scale (ACR2-SC4-01N)9. Permeate Clamp


MIDIKROS® FLOWPATH

Figure 13. MidiKros® Flowpath

LEGEND:1. Inline Polysulfone Pressure Sensor (ACPM-799-01N)2. Male Luer x 1/8” Hose Barb connector3. Manual Backpressure clamp4. Retentate Line (#14 or #16 tubing)5. 250 ml 3 port Process Reservoir (ACBT-250-C1N)6. Buffer Reservoir7. Feed Pump8. Permeate Scale (ACR2-SC4-01N)9. Permeate Clamp

MICROKROS® FLOWPATH


Figure 14. MicroKros® Flowpath

LEGEND:

1. Inline Polysulfone PressureSensor (ACPM-799-01N)

2. Female Luer x 1/16” Hose Barb

3. Manual Backpressure clamp

4. Retentate Line (#13 or #14 tubing)

5. 15 ml 3 Port Process Reservoir(ACBT-015-C1N)

6. Buffer Reservoir

7. Feed Pump

8. Permeate Scale (ACR2-SC4-01N)

9. Permeate Clamp



MidiKros® Flowpath (Figure 13, page 36)

i. Attach the pressure transducers (#1) inline to the inlet, retentate and permeate forMidiKros filters with luer connectors (X3 modules). For MidiKros filters with HB con-nections (X2), make a tee using the 1⁄8” HB plastic fittings and cut tubing.

ii. Cut an appropriate length of #16 tubing to connect from the 1⁄8” HB x HB connectoron the middle port of the 250 ml process reservoir (#5) through the pump head andthen to the luer x 1⁄8” HB fitting (#2, attached to the inline pressure transducer).

NOTE: The 50 ml reservoir (ACBT-050-C1N) can be used with #14 tubing for smaller volume

applications.

iii. Cut another length of #16 tubing (#4) from the outlet of the filter through the AutomaticBackpressure valve (if used) to the other diptube on the process reservoir.

iii. Close off the lower permeate side port of the HF module with a male Luer cap (pro-vided with module).

iv. Connect an appropriate length of tubing from the Luer x HB connector on the pres-sure sensor to the desired permeate collection vessel (not provided).

v. Plug the pressure transducer cables into the appropriate labeled jacks on the Octopuscable at the rear of the pump.

vi. Attach a buffer reservoir (#6) to the vent line (1/8” ID tubing) on the 250 ml reservoirfor constant volume diafiltration mode.

vii. If preferred, suspended tubing can be secured in the tubing notches on the sides ofthe module and reservoir Trilobites.

viii. All HB connections may be secured with tie wraps and a tie wrap gun.

MicroKros® Flowpath (Figure 14, page 37)

i. Attach the pressure transducers (#1) inline to the inlet, retentate and permeate luerports. For small volume applications (~1ml holdup volume), the pressure transducerscan be put on tees using the 1⁄16" Tees, cut tubing and 1⁄16" HB x Female Luer connectors

ii. Cut an appropriate length of #14 or #13 tubing to connect from the 1⁄16” HB x HB con-nector on the middle port of the 15 ml process reservoir (#5) through the pump headand then to the luer x 1⁄16” HB fitting (#2, attached to the inline pressure transducer).

NOTE: The 50 ml reservoir (ACBT-050-C1N) can be used with #14 tubing for large volume

applications.

iii. Cut another length of #14 or #13 tubing (#4) from the outlet of the filter through theAutomatic Backpressure valve (if used) to the other diptube on the process reservoir.

iii. Close off the lower permeate side port of the HF module with a male Luer cap (provided with module).

iv. Connect an appropriate length of tubing from the luer x HB connector on the pressuresensor to the desired permeate collection vessel (not provided).

v. Plug the pressure transducer cables into the appropriate labeled jacks on the Octopuscable at the rear of the pump.

INTEGRITY TESTING


vi. Attach a buffer reservoir (#6) to the vent line (1⁄32” ID tubing) on the 15 ml reservoir for con-stant volume diafiltration mode.

vii. If preferred, suspended tubing can be secured in the tubing notches on the sides of themodule and reservoir Trilobites.

viii. All HB connections may be secured with tie wraps and a tie wrap gun.

10.2.2 Batch ModeFor batch mode, there is no buffer reservoir connected to the vent line. Batch mode is typically usedfor concentration or clarification of a sample that is able to fit in the selected reservoir.

10.3 Integrity TestingAll Spectrum HF membrane modules are integrity tested in our class 10,000 cleanroom prior to pack-aging and shipping. Membrane integrity tests generally measure the capillary pressure of wettedpores. It is strongly recommended to integrity test all HF membrane modules again prior to use. Drymodules must be fully primed to ensure complete membrane wetting, otherwise a false integrity fail-ure will be indicated due to non-wetted or incompletely wetted pores. For wetting and integrity test-ing procedures, please refer to the “Hollow Fiber Module – Preparation and Instruction Guide”(Document 400-12058-000) provided with each hollow fiber module. For specific instructions on howto use the KrosFlo® Research IIi System to wet out the module and to perform an integrity test, seebelow.

10.3.1 Leak TestThe flowpath connections should first be pressure tested with air prior to filling the systems with fluids.

1. Close the pinch clamp on the permeate line. Close the backpressure control valve on theretentate line.

2. Turn on the pressure monitor and ensure that it is tared to zero.

3. Open the vent on the processing reservoir and run the pump until a pressure of ~5 psi isreached.

4. If there is no significant pressure drop (<0.5 psi/min), the system has no leaks and isready for use.

5. If the pressure decay is >0.5 psi/min, then check to make sure all the seals and fittingsare secure. Adjust connections as necessary to eliminate pressure decay.

6. After assuring system is free of air leaks, relieve the pressure by opening the backpres-sure clamp.

10.3.2 Wetting the HF Membrane Module

1. Fill the process reservoir with the appropriate wetting agent: 30 - 70% IPA or EtOH fordry polysulfone membranes, or DI water for the other membrane types (prewetted poly-sulfone, mixed cellulose ester or polyethersulfone).

NOTE: Rinsing the module can also be done offline to avoid contaminating the flowpath when


INTEGRITY TESTING

alcohol is used. Use a proper sized beaker or similar container as the feed and return.

2. Turn the pump on and increase the velocity until a flow pressure of ~3 – 5 psi is gener-ated. At this point, air pockets will be forced out of the flow-path and will collect in theprocessing reservoir, thus lowering the fluid level.

3. Open the filtrate shut-off clamp and rinse for 10 – 30 minutes. Alternatively, collect 2ml/cm2 of membrane surface area (refer to “Hollow Fiber Module – Preparation andInstruction Guide”). If wetting with alcohol, make sure to rinse out the alcohol with 2ml/cm2 of DI water after the system is drained.

4. If using the KF Comm data collection software, open the ‘Module Characteristics’ work-sheet and begin collecting data to determine the baseline NWP data. Vary the TMP toget a linear characterization of water flow versus pressures. The permeate flow (Qpermeate)and Temperature need to be manually entered. The ‘NWP’ worksheet will display agraph.

5. Before shutting off the pump, always close the shut-off clamps for both the buffer trans-fer and the filtrate lines.

NOTE: Whenever the system is filled with fluid and the pump is turned off, the shut-off clampsfor both the transfer and filtrate lines should be closed first. Because the retentate vol-ume is small, pressure without recirculation can plug the membrane. Although there isno pressure due to pumping, hydrostatic pressure can convert the HF module into dead-end mode.

6. Drain fluid from the system.

i. Close the buffer transfer and filtrate shut-off clamps.

ii. Lift the cap above the processing reservoir and turn on the pump to drive the remain-ing fluid in the flowpath tubing into the reservoir.

iii. Shut off the pump once all the fluid has emptied into the process reservoir. Empty theprocess reservoir and reconnect.

10.3.3 Pressure Hold Test

1. Wet the membranes in accordance with Section 10.3.2.

2. Close the retentate backpressure clamp, close the permeate clamp and open the venton the process reservoir.

3. If using the data collection software KF Comm, begin collecting data in the Integrity Testworksheet. This will automatically calculate the pressure drop in psi/min.

4. Use the pump to generate pressure in the filter until ~5 psi registers on the inlet pressuretransducer. Do not exceed 10 psig. Open the filtrate clamp and the pressure in the sys-tem will decrease as the remaining fluid is pushed through the fibers. Turn on the pumpagain and bring the pressure up to ~5psi. Verify that the pressure does not decay morethan 0.5 psi/min. If it does, then there is a leak in the system set-up. Tighten all seals,clamps and connections and try again.

5. If the pressure drops rapidly, there may be one or more broken fibers. If the pressuredrops slowly but >0.5 psi/min, there is either a broken fiber, a pin-hole leak or an incom-pletely wetted membrane. Try wetting the module again according to Section 10.3.2 andthen perform another Pressure Hold Test. If the pressure drops faster than 0.5 psi/min

INTEGRITY TESTING


again, use a different membrane module.

10.4 Modes of Operation

The basic operating modes for the KrosFlo® Research IIi System are:

• Batch Concentration of cells, virus, precipitates, proteins or diagnostic particles

• Batch Clarification of cells, cell debris, virus, precipitates or proteins

• Topped-off Batch for reduced volume operations

• Diafiltration (washing) of cells, cell debris, virus, precipitates, proteins or diagnostic particles

10.4.1 Batch Concentration (product in the retentate)Generally, the term “concentration” is applied to applications where the material retained by the mem-brane is (or contains) the desired product. As the process fluid is recirculated through the membraneand back to the feed reservoir, the feed reservoir becomes the process reservoir — the volume dimin-ishes due to the removal of permeate and the product is concentrated. This mode of operation isused, for example, with fermentation recoveries in which the desired product is the cell itself or as aninitial processing step in which the product is intracellular.

10.4.2 Batch Clarification (product in the permeate)The term clarification is generally used for applications in which the desired product is in the perme-ate, such as soluble proteins. This mode of operation is used, for example, to harvest animal cell cul-tures in which desired product is secreted by the cells, or microbial fermentation in which the desiredproduct has been released into solution by cell lysis.

When operated in concentration or clarification modes, the membrane quantitatively removes solidslarger than the pores of the membrane and allows the passage of soluble materials that are smallerthan the membrane pores. Set-up is the same for both batch concentration and clarification process-es. The retentate is returned to the feed reservoir, acting as a process reservoir, and more clarifiedproduct permeates through the membrane. The degree of concentration, called the concentrationfactor (CF) or volume reduction factor (VRF), is given by the following equation where Vi is the initialvolume and Vf is the final volume:

CF = VRF = Vi / Vf.

10.4.3 Constant VolumeA disadvantage of cross-flow batch operations is the relatively high flow rate required for efficient tan-gential flow — which is 20 – 100 times the permeate rate. As a result, it is difficult to achieve high con-centration factors without foam or vortex formation in the process reservoir. A way of avoiding thisproblem is to operate the membrane in constant volume mode, where a smaller intermediate reser-voir is utilized as the process reservoir to maintain a constant “working” volume. While the pump cre-ates a positive pressure that drives the filtration, it also creates an equal negative pressure (vacuum)that pulls the feed in at the same rate. So there is an overall volume movement from the buffer addi-tion reservoir through the process reservoir and out the filtrate line. The module simply inhibits thepassing of any material larger than the membrane pore size and concentrates it in the working vol-ume in the process reservoir.

In the example of harvesting from a fermenter or stirred-tank bioreactor, attempts are often made touse the bioreactor vessel as the process reservoir with poor results. The fermenter exit and entranceports are frequently too small for adequate recirculation flow. The return line must be modified so that


INTEGRITY TESTING

it is submerged at high concentrations (low volumes). Foaming or vortexing prevents achieving thedesired concentration factor, and results in a relatively large amount of unprocessed broth. More rapidand higher concentration can be achieved by using a secondary process reservoir and operating in aconstant volume mode.

10.4.4 Diafiltration Materials that pass through the membrane can be washed away from materials that are retained bythe membrane (cells, particles, etc.). This technique, called diafiltration, is used to recover additionalproduct in clarification applications with the goal of achieving better product purity in concentrationapplications. For best efficiency, the wash buffer should be free of the solute that is being recoveredor removed. Diafiltration may be accomplished either by adding buffer at the same rate as the per-meation rate (constant volume diafiltration) or by reducing the volume in the feed reservoir and addingmore buffer to regain the original volume (discontinuous diafiltration). The amount of diafiltration per-formed can be expressed by the volume of wash buffer added divided by the batch volume, i.e. thenumber of “wash volumes”. During a constant volume diafiltration, in which soluble components passfreely through the membrane, each wash volume of permeate removed reduces the solute concen-tration by a factor of e (2.718…). For example, a four time wash volume diafiltration will reduce theconcentration of solute by a factor of e4, i.e. 50-fold or over 98%. Using this technique, the concen-tration of solute can be monitored in the permeate until the desired level of purification of productrecovery is achieved. Components that are partially retained by the membrane cannot be diafilteredor washed with the same efficiency. The membrane rejection (or retention) is defined as:

Rejection = 1 – Concentration in the permeateConcentration in the retentate

As the membrane rejection of an undesired component increases, diafiltration becomes less effectiveas a technique for removing it.

Figure 15. The Effect of Solute Rejection on Diafiltration Efficiency

The system set-up for diafiltration is the same as for topped-off batch mode, except that washbuffer enters the process reservoir instead of more product. For more detailed information on diafil-tration, please refer to the diafiltration literature on Spectrum Labs’ website: http://www.spectrumlabs.com/lit/hfdial.pdf.

OPERATING THE SYSTEM



OPERATING THE SYSTEM

10.5 Operating the SystemThere are two basic system set-ups and operating instructions for the different modes. “Batch” oper-ating instructions are used when the KrosFlo® Research IIi TFF System is used for batch clarificationand batch concentration. “Constant Volume” operating instructions are used when the system is usedfor topped-off batch and diafiltration modes.

10.5.1 Instructions for Batch Concentration / Clarification

1. Assemble the system according to the instructions in Section 10.2.2.

2. Integrity Test: Test the integrity of the system and the membrane module according tothe instructions in Section 10.3.

3. Introduce the sample to be filtered into the reservoir and seal the lid.

4. Priming: Close the permeate shut-off clamp and make sure the second permeate port(drain) is closed. Turn the pump on in the forward direction and slowly increase the ratejust high enough to prime the system, but low enough to avoid foaming and vortexingthe solution. Continue until a clear, bubble-free stream is coming out through the reten-tate line back to the feed reservoir. It is important to eliminate all air from the recirculationloop to avoid product shear.

5. Setting Operating Parameters: Adjust the pump rate to the desired velocity after the airis eliminated. An easy and quick guide to use is given in the header of the KF Commworksheet. Type the model number of the filter or select the filter from the pull downmenu. The recommended feed flow rates at specific shear rates are given on the rightside of the header. For fouling (eg. protein) applications or cell applications, shear ratesof 12,000 s-1 or a maximum shear rate of 4,000 s-1 are recommended respectively.

6. If using the KF Comm data acquisition software, open the ‘Trial Data’ worksheet andbegin collecting data. The permeate flow rate (Qpermeate) or total volume of permeate(Vpermeate) will need to be manually entered (unless using the optional Permeate Scale) attheir respective times to get graphs of the Flux Rate decay and the Pressures vs.Volumetric Throughput (VT). OR, equivalently, the mass of the permeate (Mpermeate) can bemanually entered at the respective times using an appropriate scale (not provided).

7. Check for system leaks and tighten fittings and connections if necessary.

8. Open the permeate shut-off clamp. Filtrate should start coming out the permeate line.

9. Adjust the pressure if necessary by using the Automatic Backpressure Valve or the flowrestrictor on the retentate line to induce back pressure.

10. Concentration / Clarification: Allow the system to run in batch mode while monitoring thefiltrate (flux) rate until the desired concentration factor has been achieved or the desiredvolume has been filtered through the membrane module. In batch mode, the flux ratemay diminish as the retentate material concentrates in the process reservoir.

11. If the retentate starts to vortex or foam, stop processing the batch. For clarification appli-cations, the system can run dry. When the batch processing is complete, close the per-meate shut-off clamp and turn off the pump.

12. Product Recovery: To recover retentate, lift the lid above the feed reservoir and turn thepump on at a low rate — this will drive the retentate in the recirculation loop into the feedreservoir using air.

13. To recover filtrate in the extracapillary space of the membrane module, open the perme-ate shut-off clamp as well as the drain shut-off clamp on the module lower side-port.Drain filtrate into the filtrate reservoir used during the filtration process.

PRE-STRAINING


14. Post-process Integrity Test: Since Spectrum hollow fiber modules are disposable, assur-ance of membrane integrity can be achieved by performing a post-process integrity test.

NOTE: A post-process integrity test renders the module non-reusable.

10.5.2 Instructions for Constant Volume & Diafiltration

1. Assemble the system according to the instructions in Section 10.2.1.

2. Integrity Test: Test the integrity of the system and the membrane module according tothe instructions in Section 10.3.

3. Fill the processing reservoir with the process fluid to be washed.

4. Establish a pump velocity (corresponding to suggested recirculation rates based on adesired shear).

5. The membrane inlet pressure and velocity of process fluid should be adjusted for eachspecific application. Transmembrane pressure can be increased by increasing the recir-culation rate, pressurizing the buffer addition tank, or pulling a vacuum on the filtrate line.

6. Membrane inlet pressure should not exceed 30 psi, the maximum membrane pressure.The KrosFlo® Research IIi System has a programmable high pressure warning alarm anda programmable high pressure pump shut-off that can be used to ensure that pressuredoes not exceed a user-defined limit.

7. Re-adjust the processing reservoir volume to a desired level by introducing more samplefrom the buffer addition reservoir, as described in Step 3.

NOTE: The level of fluid in the processing reservoir plus the volume in the tubing in the pro-

cessing loop will determine the final volume of the sample after diafiltration. Below are

the procedures to adjust the fluid level of the processing reservoir:

Lowering Processing Reservior Fluid Level

• During the filtration run, close the buffer addition shut-off clamp

• Make sure the filtrate clamp is open

• Open the vent on the processing reservoir by removing the buffer addition line

• When desired reservoir volume is reached, reattach the buffer addition line and open the shut-off clamp; to avoid vortexing and bubbling, keep the fluid level in the processing reservoir above the dip tubes that extend toward the bottom of the reservoir

RAISING PROCESSING RESERVOIR FLUID LEVEL

• During the filtration run, make sure the buffer addition shut-off clamp is open and the filtrate line is closed

• Open the cap on the recirculation reservoir and gravity feed in the buffer/sample from the buffer reservoir

• When desired reservoir volume is reached, close the cap on the recirculation reservoir and open the filtrate line

Whenever the processing reservoir volume drains with the vent closed or the buffer line

attached, then there is a leak in the system.

8. Open the shut-off clamps for both the buffer addition and filtrate lines


PROCESS OPTIMIZATION

9. Use the KF Comm data acquisition software to maintain a record of the operating con-ditions

10. After the desired number of washings has been achieved, the system can be drainedaccording to the instructions in Section 10.3.2.9

10.6 Pre-StrainingSome process streams may require a strainer to prevent the lumen of the hollow fibers from becom-ing occluded by solution components larger than the diameter of the hollow fibers. If the fiber lumensbecome blocked, the pressure between the pump and the module will increase and the permeaterate will decline. In extreme cases, the resulting pressure build-up between the pump and the inlet tothe module can result in membrane module failure. A strainer should be used on the fluid prior toentering the process reservoir.

For applications where large particles or agglomerates are a concern, a test should be performedprior to processing. Install a 50 mesh screen in place of the membrane module. Circulate the processfluid at a high rate. If the feed pressure begins to increase while the pumping rate is constant, thescreen is being blocked. If this is the case, the process fluid must be strained prior to entering theprocess reservoir.

Strain the process fluid using a 50 mesh stainless steel screen. If the pressure remains constant afterpre-straining the feed, the pre-strainer worked. If there is difficulty in pre-straining the process fluid, alarger mesh strainer can be used in tandem with modules having 1 mm fiber diameters. There are,however, some fluids that plug screens quickly but do not adversely affect the membrane.

10.7 Process OptimizationAs a pressure-driven process, the two most important variables for TFF are recirculation velocity andTMP. In terms of flux and passage efficiency, these two variables are at odds with one another. Higherrecirculation rates, which generate the convective forces parallel to the membrane surface, act tocleanse the membrane. Conversely, TMP, which generates the driving force for permeation, acts toform a cake layer. Optimum operation of the module depends on establishing an equilibrium betweenthese two parameters. Optimization requires that the pressure drop through the module be moni-tored. This pressure drop (∆P) is generated by the fluid resistance of the membrane channels and isdefined by the equation:

∆P = Pin - PoutPressure drop is generated by the recirculation rate through the module and is related to the charac-teristics of the fluid, such as viscosity and density, as well as the characteristics of the module, suchas membrane diameter and length. The TMP is the average pressure differential generated across themembranes and is defined by the equation:

TMP = (Pin - Pout) / 2 – PpermeateTMP is generated by the recirculation rate though the module plus any downstream restrictions. Thefollowing method produces a graph of Steady State Flux vs. TMP that generally resembles Figure 16(see next page):

IN-PROCESS MODULE CLEANING


Figure 16. Typical Flux and Solute Passage Curve

The first section of the curve (1) is the pressure dependent region and indicates that a cake layer hasnot yet been formed. In the pressure dependent range, solute passage is generally unrestricted —raising the TMP increases the flux. The top section (2) of the curve is the pressure independent rangeand indicates that a cake layer has been formed. In the pressure independent range, solute passageis controlled by the cake layer. Raising the TMP does not increase the flux. The mid-section of thecurve (3) shows the onset of cake layer formation. The optimum TMP is the highest flux where solutepassage is close to 100%. The following is a step-wise approach to optimizing the operating condi-tions:

1. With the permeate line clamped off and no downstream restriction, begin the run with thehighest possible recirculation rate. This may be dictated by one or more considerations:inlet pressure to the module, pump capacity, solution shear sensitivity, etc.

2. Open the permeate line. Allow the permeate rate to reach a steady state, which normal-ly takes 5 – 10 minutes. Measure the flux and take a permeate sample to determinesolute passage. Collect the data (recirculation rate, transmembrane pressure, and flux)using KF Comm on a computer. Collect the solute passage data separately.

3 Increase the TMP of the module by adding backpressure. This can be done using aretentate backpressure valve, or by applying fluid or head pressure to the process ves-sel. Generally this increase should be in 0.5 bar increments. If the module was alreadybeing run at the maximum inlet pressure of 2 bars (30 psig), then the initial recirculationrate will have to be decreased.

4. Establish a new equilibrium, measure the flux, take another sample of the filtrate, andrecord the recirculation rate and transmembrane pressure.

5. Repeat Steps 3 and 4 until either flux does not increase, or until the passage of desiredsolutes begins to decline. Optimum is just prior to the point of flux decline and decline ofsolute passage.

In certain instances, even the highest recirculation rates and lowest TMP’s may generate caking. Inthese cases, restricting or metering the permeate will reduce TMP by increasing the permeate pres-sure (refer to the TMP equation). When permeate pressure is other than atmospheric, the pressuretransducer must be connected to the permeate line. In Step 2 above, partially open the permeate line,then proceed to Step 3.

TMP is increased in this case by opening the permeate line or speeding the metering pump incre-mentally and checking the permeate for solute passage. Continue to increase TMP in this manner


MODULE SELECTION AND SCALE-UP

until solute passage declines. Optimum is just prior to the point of flux decline.

10.8 In-Process Module CleaningEven under optimum conditions, a slow decrease in the permeate rate may occur. There are threetechniques for restoring permeate flux. These include: pump off, forward flushing, and reverse flush-ing. All three, however, stop or reverse permeation (which must be considered in evaluating the ben-efit of these techniques).

10.8.1 Pump Off CleaningTurning the pump off will allow the cake layer to loosen. Restarting the pump will flush loosened mate-rial from the membrane lumen surface. Although allowing one minute before restarting the pump isusually sufficient, the optimal time will vary with the solution being processed and should be investi-gated for each individual case.

10.8.2 Forward FlushingClosing the permeate shut-off clamp while recirculating will clean the membrane in the downstream-half of the module. Closing the permeate line causes the permeate pressure to rise and exceed theretentate pressure in the downstream-half of the module. In this region, the permeate will flow fromthe exterior of the fiber to the interior — loosening and carrying away caked material. Normally, for-ward flushing for one minute is sufficient to clean the downstream-half of the membrane. Open thepermeate line again and continue processing.

The principle of forward flushing can be explained as follows: when the permeate line is closed, thenet filtration rate in the module is zero — however permeation still occurs internally. The inlet-half ofthe membrane module (the high pressure end) generates permeate that back flushes the down-stream-half of the membrane module (the low pressure end). This phenomenon is called SterlingFlow.

Reversing the pump with the permeate line closed will back flush the other half of the module and isan effective cleaning technique.

10.8.3 Reverse FlushingTo clean both halves of the module simultaneously, operate the pump in the reverse direction suchthat the recirculation flow enters the module from the retentate end of the module and exits the inletend. Open the permeate line and allow the permeate to be pulled back into the module through thepermeate port for one minute. Then switch the pump to the forward direction again and continue pro-cessing.

Reversing the pump induces a negative pressure along the entire length of the membrane fibers andcauses permeate to be pulled back through the walls of the membrane and into the retentate. Thisflow reversal will flush caked material back into the bulk process fluid. Using caution, the membranecan also be back flushed by pumping permeate directly into the extracapillary space of the module.This must be done with caution at 0.7 bars differential (10 psid) maximum or the membrane hollowfibers can be permanently damaged.

Note that reverse flushing will cause the bulk process fluid volume to increase in the processing reser-voir. Precautions must be taken to accommodate for this volume increase.

10.9 Module Selection and Scale-up

The KrosFlo® Research IIi TFF System can successfully be used to determine the membrane surfacearea needed for scale-up. The performance of the membrane module depends on many processing

MODULE SELECTION AND SCALE-UP


variables; such as the fluid type being processed, the conditions of the fluid (temperature, % solids,viscosity, etc.), fluid volume, recirculation rate, extent of caking, etc. For scale-up purposes, theamount of surface area and module size needed to process a batch depends on the following:

• Steady state flux

• Batch size to be processed

• Desired process duration

• Time required for equipment turn-around (cleaning, rinsing, etc.)

Steady state flux can usually be determined by a trial run with the fluid to be processed. The batchsize and the desired time to process the batch are determined by the user. Spectrum KrosFlo® mod-ules are especially useful for small volume scale-up studies.

It is generally prudent to include a safety factor in designing the process. Oversizing the membranesurface area by 10 – 30% is generally sufficient to account for problems associated with fluid vari-ability.

In order to determine the amount of membrane surface area required for scale-up, the steady stateflux must be determined. Accuracy of the scale-up procedure is dependent on several variables beingconstant from the test runs to the final process. These variables include: solution composition, recir-culation rate, processing temperature and operating pressures. Probably the most important consid-eration is that the solution being tested is representative of the scale up fluid to be processed. Steadystate flux occurs when the permeate rate remains essentially constant over a period of 15 to 30 min-utes.

By knowing the steady state flux, the batch size to be processed and the desired processing dura-tion, a simple calculation can be made to determine the necessary membrane surface area:

Required surface area = Filtrate volume desired Required time x Steady state flux

Units are:

Required surface area = square meters (1 m2 = 10,000 cm2)

Filtrate volume desired = liters

Required time = hours

Steady state flux = liters per square meter hour (L/m2hr)

Example 1:

Clarify animals cells from 1.5 liters of bioreactor broth

The steady state flux is 90 L/m2hr.

The desired processing time is 45 minutes or 0.75 hours.

Required Surface Area = 1.5 liters = 0.022 m2 or 220 cm2

0.75 hours x 90 L/m2hr

Referring to Spectrum’s product catalog, part no. M52M-260-01N, a MiniKros® Sampler Plus mod-ule with 460 cm2 is sufficient for this application and will actually process 1.5 liters of permeate in 0.36hours (22 minutes).


PERMEATE SCALE

11. Optional Equipment

11.1 Permeate Scales Compatible with the System KrosFlo® Research IIiSystem

11.1.1 Introduction

The KrosFlo® Research IIi TFF System is the ideal tool for performing small scale Tangential FlowFiltration processes. Permeate scales compatable with the KrosFlo® Research IIi System, such as theKrosFlo® Research IIi Permeate Scale and the Ohaus® Navigator XL Scale, are accessory compo-nents that automate the data collection by measuring the mass of the collected permeate over time.Both scales connect to the KrosFlo® Research IIi System via serial connection. The system sends thedata to an Excel® Spreadsheet through a USB connection. The KF COMM software then automati-cally tabulates and graphs the data.

NOTE: All editions of KF COMM prior to 1.14 do not come with the Ohaus® Nevigator scale driver

preloaded.

PERMEATE SCALE


11.1.2 Installation

1. Connect Permeate Scale serial cable to “SCALE” serial port on Pump Drive’s Octopus cable,then connect Permeate Scale to power source

2. Attach USB extension cable to the USB connector labeled “COMPUTER” on the OctopusCable, and plug in the other end of the USB extension cable to the PC

3. Turn on Pump Drive, but leave Permeate Scale off

NOTE: If on, Permeate Scale will not be recognized by PC when booting.

4. Boot PC, and allow Windows® to search for the Pump Drive’s driver — if not found, locatethe driver on the included CD when the prompt displays

5. Turn on Permeate Scale

6. Open Excell® Workbook “Scale Data.xlt”

7. Enable Workbook’s macros (this may not be possible if user’s security settings are not highenough), if successful:

a. Older versions of Excel® will display four icons for data collection control on the standardtoolbar

b. Newer versions of Excel will display data collection controls in Add-Ins tab

8. Establish and verify Pump Drive’s built-in Pressure Monitor connection

a. Click on “Configure Collection” to set up Pressure Monitor


Figure 17: Configure Scale Menu on Windows XP

i. Locate Pressure Monitor by clicking “Find COMM,” which will prompt the PC to contin-uously cycle through available COM ports until it connects to Pressure Monitor, or untiluser clicks “Cancel”

ii. If the Pressure Monitor’s COM is unavailable due to another program’s interference,close interfering program

b. Verify Pressure Monitor’s live connection to PC by adding pressure to Inline PressureSensor — the “Trial Data” spreadsheet will update its corresponding yellow Live cell,depending on which port Pressure Sensor is connected to in Octopus Cable, at top ofthe sheet to reflect the pressure

NOTE: “Override.xls” spreadsheet may be used to force Pressure Monitor’s COM port to the cor-rect setting.

9. Establish and verify Permeate Scale connection

a. Click on “Configure Scale” to set up Permeate Scale

Figure 18: Configure Permeate Scale Menu on Windows 8

!

PERMEATE SCALE

PERMEATE SCALE


i. Locate Permeate Scale by clicking “Find Comm,” which will prompt the PC to continu-ously cycle through available COM ports until it connects to Permeate Scale, or untiluser clicks “Cancel”

ii. If Permeate Scale’s COM is unavailable due to another progran’s interference, closeinterfering program

iii. Select “Scout Pro” under “Scale Accessory for KrosFlo® Research IIi TFF System” ifusing KrosFlo® Research IIi Permeate Scale, or “Navigator” if using Ohaus®

Navigator XL Scale in the drop-down menu within “Configure Scale” dialog

iv. Click “OK.”

b. Verify Permeate Scale’s live connection to PC by placing a small object on the scale —the “Trial Data” spreadsheet will update its yellow Live cells at top of the sheet to reflectthe small object’s mass.

NOTE: “Scale Override.xls” Spreadsheet my be used to force Permeate Scale’s COM port to the

correct setting

10. Permeate Scale is now ready to use in conjunction with KrosFlo® Research IIi TFF Systemand will zero automatically when the “Start Collection” button is pressed

11.1.3 Troubleshooting

a. COM Port Setting

i. If the COM port cannot be detected for the Pressure Monitor or the Permeate Scale, thePressure Monitor Override or the Scale Override Excel® spreadsheets can be usedrespectively. These are provided on the CD and force the COM port to the correct set-ting. To do this, open the file and change the COM port to the correct setting (seen inthe Device Manager), then save and close the file. Reopen the spreadsheet and selectyes when asked to force the COM setting.

b. Driver Recognition

i. If Windows® does not recognize the Permeate Scale USB signal or thinks that it is a dif-ferent device, unplug the USB connector from the computer, turn off the scale and plugin the USB connector again. If the driver is still not recognized, try loading the driveragain and/or rebooting to reset the drivers.

11.1.4 Updating KRIIi Firmware

1. Connect the pump to the computer and turn it on

2. Ensure both the ProgramKRIIi.exe and the KR2iSource.cod file are in the same directory

3. Start the ProgramKRIIi.exe by double-clicking on it

4. Click on the Start button to begin the process

5. Either click the Find Comm button, or select the Comm port used by the pump; it will bethe same as the Comm port selected in one of the workbooks if you are using them.

6. Click on the Go button and the pump display will go dark and the pump will begin toupdate. This process will take approximately 10 minutes.


11.2 Automatic Backpressure Valve for KrosFlo Research IIi System

The KrosFlo® Automatic Backpressure Valve controls TMP or permeate pressure duringtangential flow filtration processes when used in conjunction with the KrosFlo® ResearchIIi TFF System. The valve is designed to pinch flexible tubing on the retentate or per-meate line based on digital pressure readings and maintain the user-set pressure.

11.2.1 Installation

AUTOMATIC BACKPRESSURE VALVE

7. To verify a successful update, restart the pump. The boot-up sequence should now dis-play the most current version.

If the pump is interrupted while updating, the display will remain dark. Restart both thepump and ProgramKRIIi.exe and proceed as before. If the display remains dark for anextended period of time while programming, restart the process. The pump will not oper-ate until the program updates successfully.

1. The cable on the KrosFlo® Automatic BackpressureValve has both an RS232 port (female) and a powersupply. Connect the RS232 port to the terminal labeled“Valve” on the Octopus cable at the back of the DigitalPressure Monitor. Ignore the power supply connection;it is used to power a stand-alone pressure monitor.

2. Connect the second RS232 port on the opposite side(male plug) to the serial cable for sending data to a PC.The data can also be sent to the PC using the“Computer” USB connection on the Octopus cable.

3. Plug in the transformer and connect to the power sup-ply on the Automatic Backpressure Valve.


11.2.2 Operation

Automatic TMP Pressure Control

1. To operate the backpressure valve in the auto TMP mode, switch the toggle on the topof the valve to the “TMP” setting. Press the OPEN button to load the tubing and resetthe pressure controls.

2. Set the desired TMP using the INC and DEC buttons

!


Power RequirementsACPC-F16-01N, 1/4” (max) OD tubing valve � 0.5A @ 12V

ACPC-F17-01N, 5/8” (max) OD tubing valve � 1.0A @ 12V

ACPC-F82-01N, 1” (max) OD tubing valve 1.0A @ 12V

KRIIi’s Port

for ABV

ABV’s Port

for KRIIi


3. With the pump running at the desired speed for the TTF process, press the GO buttonand the valve will start to pinch the tubing until the desired TMP is reached (this may takea few minutes until it equilibrates).

4. To pause the valve, press the GO button (the light will turn off). To resume, press the GO but-ton (the light will turn back on).

NOTE: Under manual control, the Automatic Backpressure Valve does not need to equilibrate.

NOTE: If the valve is not in the fully open position, it will adjust the pinch slowly to avoid the

potential pressure spikes. To manually force it to adjust faster, hold the GO button down

for >3 seconds until the lights begin to blink — indicating that the backpressure valve is

now under manual control — and then press the GO button again.

b. If the KrosFlo Research IIi Pump is turned to off or to 1/2 the rpm when the GO buttonwas pressed, the valve will pause and will maintain the same pinch distance. The valvewill flash “SLO” while in this paused state. The valve will resume the normal pinch con-trol when the pump is turned on or to at least 1/2 the original rpm.

Automatic Permeate Pressure Control

1. To operate the backpressure valve in the auto PERMEATE mode, switch the toggle onthe top of the valve to the “Permeate” setting. Press the OPEN button to load the tub-ing and reset the pressure controls.

2. Set the desired Permeate pressure using the INC and DEC buttons

3. Press the GO button before starting the recirculation pump — the valve will pinch shut(when using the maximum tubing size rated for the valve). Begin the process and thenthe valve will start to open/pinch the tubing until the desired Permeate pressure isreached (this may take a few minutes to equilibrate).

ManualTo control the valve manually, hold down the GO button for >3 seconds. The light will begin to flash and then the INC and DEC buttons can be used to adjust the pinch.

Updating Automatic Backpressure Valve1. Connect the KrosFlo® Automatic Backpressure Valve to the computer via the RS232

port

2. Turn on the valve and wait 60 seconds

3. Ensure the appropriate update has been obtained, either for the ACPC-F16-01N orACPC-F17-01N valve.

4. Press and hold the Open button on the valve for 6 seconds, blanking the display

5. Start the .exe file by double-clicking on it

6. Click on the Start button to begin the process

7. Either click the Find Comm button, or select the Comm port used by the valve

8. Click on the OK button and the vavle display will go dark — the vavle will begin to update

9. To verify a successful update, toggle the TMP/Permeate to restart the valve. The valveshould now display the most current version


Communications

Serial Communication

The Valve has a serial input cable for communication and control.

The communication parameters are 19200 baud, 1 stop bit, no parity, and no flow control.

When communicating with an external PLC, the valve should receive about 1 line of dataper second. The lines should end with a return character and a line feed character.

Each line should have several values, each separated from the next by a single space:

<InletPressure><space><RetentatePressure><space><PermeatePressure><space><TMP><space><RPM><space><return>

1. Inlet Pressure: This should contain a decimal and may have a leading - sign for negative pressures. There should be no sign provided for positive values. If the - sign is included, there should not be a space after the - sign. The number will not be less than 2 charac-ters, nor more than 5 characters in length. The number will contain a decimal point. If inunits of millibars, there will be no digits to the right of the decimal. If in psi, there will be1 or 2 digits to the right of the decimal. The pressure must be no more than 50 psi norless than -9.99 psi.

2. Retentate Pressure: Formatted same as the inlet pressure.

3. Permeate Pressure: Formatted same as the inlet pressure.

4. Transmembrane Pressure (TMP): Formatted same as the inlet pressure.

5. Pump speed in RPM: This is a number with no decimal point and no possible leading - sign. It is the unsigned pump speed in RPM. It may be 0. When the GO button ispressed, the pump speed is remembered. If the speed drops by 50% or more, the valvestops adjusting position until the pump speed recovers past the 50% mark. If the RPMis 0 when the GO button is pressed, then the pump speed will never fall below the 50%limit and the Pump Slow mode will never be entered.

Note: The pump speed should be followed by a space. This space may be followed by other characters, but they are ignored by the valve.

The line should be no more than 78 characters in length.

Valve External Control

If the input line is “G0,” then the valve will stop and the green light will turn off.If the input line is “G1,” then the valve will go and the green light will turn on.If the input line is “G2,” then the valve will open for loading or unloading tubing.

Please note that while the valve is moving a long distance (as it usually doesimmediately following a go or load command), additional commands are silently ignored.

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Also, the set point may be adjusted from the data stream. Use "S # " the letter S followed by a space, followed by a number with a decimal point, followed by a space, which may be followed by any additional characters desired. The number should conform to the stan-dards used for the pressure readings (no decimal digits is mb, 1 or 2 decimal digits is psi, and 3 decimal digits is bar).

The S and G lines count toward the valve timing. Wait at least 100 ms between sending lines.

Fine Tuning

The valve positioning and timing can be modified using terminal emulators, such as HyperTerminal (available on Mircrosoft® Windows 95 up to XP) or PuTTY (a free and open-source program for multipul platforms).

To set the start position, connect the valve to the computer using a serial cable (or USB to serial adapter if no serial connector is available on the computer). Do not plug the serial connector into the pressure monitor. Turn on the valve and wait 60 seconds. Press and hold the OPEN button for about 6 seconds until the display is all - signs. Use the terminal emulatorl to communicate with the valve, using the usual settings — 19200baud, 8 data bits, no parity, 1 stop bit, and no flow control. Select the correct COM port and make sure that the KF Comm software is not running.

Once connected, press the enter key and then type:<Start><space><start position number>

Example: Start 98 and then the enter key to set the start position. The default number is usually 80. Larger numbers are more closed. Numbers bigger than 255 are reduced to numbers less than 256.

To display the current start position, type <Start> and press enter. When the correctstart position number has been entered, type <Lock> and press enter to go back tonormal valve operation. The number will be stored, and the valve will close to thenew position when GO is pressed.

There are 4 other parameters that can be tuned if needed:

1. “Change” is the minimum number of seconds before a change is made in the positionafter a change in the sign of the pressure error. The default is 3.

2. “Interval” is the minimum number of seconds between changes in position. The defaultis 4.

3. “Step” is the number of steps to take in the same direction before the movements getlarger. The default is 5.

4. “Reverse” is the minimum number of seconds to wait after the first movement in a newdirection. The default is 9.

12. Ordering InformationKrosFlo® Research IIi Tangential Flow Filtration System

Part Number Description

SYR2-U20-01N KrosFlo Research IIi System with pump drive, pump head and

Integrated Digital Pressure Monitor.

KF Comm software, Octopus cable, Flowpath kit and Starter kit also included.

ACR2-IQOQ-PW KrosFlo Research IIi System IQ/OQ Paperwork

ACR2-IQOQ-SR KrosFlo Research IIi System IQ/OQ Sales Rep Service


ORDERING INFORMATION

System Components & Accessories

Part Number DescriptionACPM-201-01N KrosFlo Pressure Monitor, 110V; w/ KF Comm, 3 transducers & RS232

ACPM-202-01N KrosFlo Pressure Monitor, 220V; w/ KF Comm, 3 Transducers & RS232

ACPM-499-03N Disposable Pressure Transducer, Sterile, 3/Pkg

ACPM-799-01N Polysulfone Pressure Transducer 1/Pkg

ACPM-799-01S Polysulfone Pressure Transducer, Irradiated 1/Pkg

ACPX-CD-01N MicroKros/MidiKros Fitting Kit

ACPX-T-01N MidiKros TC Fitting Kit

ACPX-S-01N MiniKros Sampler Fitting Kit

ACR2-SKT-01N KrosFlo Research IIi Starter Kit

ACR2-UFP-01N Universal Disposable Flowpath Kit for use with L/S 13 To L/S 17 Tubing

ACR2-H3I-01N KrosFlo Research IIi Pump Head, 3 Stainless Steel Rollers

ACR2-H4I-01N EZ Load 3 Pump Head for Thick Wall Tubing w/ Interlock

ACPC-F16-01N KrosFlo Automatic Backpressure Valve for use with 1/8" To 1/4" OD Tubing, 110V.

(L/S # 13, 14, 16 Tubing)

ACPC-F17-01N KrosFlo Automatic Backpressure Valve for use with 3/16" To 5/8" OD Tubing, 110V.

(L/S # 25, 17, 18, 15, 24, 35, 36 and IP # 26, 73)

ACPC-F82-01N KrosFlow Automatic Backpressure Valve for use with 3/4” to 1” OD Tubing, 110V.

(L/S # 73 and #82 Tubing)

ACR2-SC4-01N KrosFlo Research IIi Permeate Scale 4000g (0,1 g), 110V




ACR2-SC8-01N Ohaus®

Navigator Scale 10000g (1.0g), 110V







ACBT-015-C1N Process Reservoir-Conical Bottom, 15 ml, 3 Port




ACBT-1HB-01N Process Reservoir, 1L, Polypropylene, Mini HB connections

ACBT-1TC-01N Process Reservoir, 1L, Polypropylene, 3/4" TC Connections

ACBT-2HB-01N Process Reservoir, 2L, Polypropylene, Mini HB connections

ACBT-2TC-01N Process Reservoir, 2L, Polypropylene, 3/4" TC Connections

ACTO-4TC-01N Process Reservoir, 4L, Polypropylene, Mini TC Connections

ACTO-2PP-01N Process Reservoir, 2L, Polypropylene, two dip tubes, 1 vent

ACTO-2PC-01N Process Reservoir, 2L, Polycarbonate, 1/4" HB connections

ACTU-P13-25N Pharmapure® Tubing, L/S 13, (1/32" ID), 25 ft/pkg






ACTU-E13-25N Extended Life Silicone Tubing Size 13, 0.03" (0.8mm) ID, 1/16" Hose Barb





NOTE: Contact Spectrum for assistance selecting the appropriate HF Module, Flowpath, Reservoir Bottles and Customized Sterile Flowpaths (MBT's) for your application.

Sensor Monitor

Fitting Kit

Pump Head

ABV

Scale

Tubing

Reservoir


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