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Sartobind Capsules for Economic Production of Therapeutic Proteins
High speed polishing
Sartobind SingleSep® capsules are designed
to remove charged contaminants from
therapeutic proteins at accelerated flow
rates by ion exchange membrane chro-
matography. The high throughput is a
direct result of negligible mass transfer
effects and is made possible by the >3 μm
macroporous membrane with 4 mm bed
height.
Approved technology
In 2001 Campath®* received FDA approval.
This monoclonal antibody is polished by
application of Sartobind ion exchange
Q modules in flow through mode (DNA|
endotoxin|leached protein A|virus removal).
It was the first time that a Membrane
Adsorber has been accepted in the produc-
tion of a therapeutic protein. This shows not
only Sartorius’ position as the pioneer in
adsorptive membrane chromatography but
also displays that this technology is proven
and tested.
* Campath is a trademark of Genzyme
Corporation
Efficient
– Higher throughput (g/h) for trace
impurity removal
Economical
– Saves capital
– No hardware investment & maintenance
– No column packing, testing,
regeneration
– No re-use validation
– Less unspecific binding – higher yield
– Less labor
– Buffer consumption may be
decreased 95%
Easy to use
– Disposable
– Simple and fast set up
– Handling like a filter capsule
Controlled quality
Sartobind SingleSep capsules are designed,
developed and manufactured in accor-
dance with an ISO 9001 certified Quality
Management System. They have passed
Plastic Class VI, particles and extractables
test according to current United States
Pharmacopoeia (USP) and are tested for
protein binding capacity prior to release.
Each package contains a certificate of
quality.
A validation guide is available upon
request: Order No.: 85032-536-72.
Ion Exchange Membranes
Sartobind Q
The workhorse to remove negatively
charged contaminants: DNA, host cell
proteins, viruses and endotoxins.
Sartobind S
For the removal of positively charged
contaminants such as host cell proteins.
Sartobind STIC®
In addition to conventional anion exchange
chemistries, Sartobind STIC PA (primary
amine) ligand operates at high salt condi-
tions to decrease need for dilution (e.g. of
cation exchange pools) when removing
negatively charged impurities. With a
binding capacity of 1.4 mg/cm2 the salt
tolerant adsorber is a specialist for (host
cell) protein removal. MVM virus removal
at 150 mM NaCl salt result in >4 LRV and
DNA removal in >3 LRV.
Capsule design
The Sartobind SingleSep looks like a
standard filter capsule except that the
adsorbing membrane is reeled on a core
to form a cylinder. The flow is from top
into the outside channel and then perpen-
dicular through the membrane layers to
the center of the cylinder and leaves the
capsule at the outlet.
3
l Fig. 1: Sartobind 30 inch capsule connected to a disposable bag.
Materials
Labour
Consumables
Hardware costs
IEX gel column
MaterialsLabourConsumables
�
80% less
IEX Membrane Adsorber Capsule
Flow path/4 mm bed height
Sartobind capsule
Fig. 2: Cost comparison of columns and Membrane Adsorbers for polishing application
Fig. 3: Schematic flow pattern and cross section of capsulesa) Sartobind pico 0.08 ml b) 10" capsule 180 ml
a)
b)
Application Area
In a generic downstream process for the
production of a monoclonal antibody, the
main capturing steps are followed by the
polishing step(s).
The later the anion exchanger is applied in
the process, the less contaminants have to
be removed.
The positively charged ligand of the
Q adsorbers binds negatively charged
contaminants.
Neutral or positively charged product
runs through (see Figure).
+
_+
4
Cell removal|Clarification
Affinity Chromatography
Cation Exchange Chromatography
Anion Exchange Adsorber Polishing Step (flowthrough):
Removal of DNA, HCP, Endotoxins
Virus Clearance
Ultra-|Diafiltration
Sartobind usage in monoclonal antibody production:
Endotoxin
DNAVirus
Host cell protein
2 7 12
pH of buffer
MAb
_ _
_
_
_
+
+
pl
Fig. 4: Isoelectric points and charges of MAbs and impurities
Binding capacity on Sartobind ion exchanger membranes
IEX ligand Type of molecule Binding capacity Binding capacity Ref.
chemistry per ml per cm2
Q BSA >29 mg >0.8 mg 1
STIC PA BSA 50 mg 1.4 mg 4
Q DNA 5.6 mg 0.16 mg 2
STIC PA DNA 10 mg 0.3 mg 4
Q Adenovirus up to 1.5+1013 up to 8+1011 Viral Particles 3
S Lysozyme >29 mg >0.8 mg 1
1 ml = 36 cm2 IEX membrane1 Sartorius quality control for minimum static
binding capacity2 J. K. Walter, (Boehringer-Ingelheim Pharma) in:
Bioseparation and Bioprocessing, G. Subramanian(ed.) Wiley VCH, 1998, Vol. II p. 447–460
3 Measured with Vivapure AdenoPack 500. Order no.:VS-AVPQ501
4 Measured in high salt buffer, 150 mM NaCl,Sartobind STIC PA manual, Order no.: 85032-536-77
5
The use of conventional chromatography
columns for flow through (FT) anion
exchange chromatography requires high
flow rates. Optimized production columns
need a certain diameter and bed height to
achieve a large throughput and therefore
have a large bed volume. That is the reason
why columns are typically oversized.
Due to the cylindrical construction of the
adsorber the high flow rate of the
membrane is combined with a large frontal
surface resulting in very high throughput
and keeping the bed volume small. Typical
process data show that operation time can
be reduced by about 80% and buffer
reduction can reach 95%. Protein loading
in FT can be two orders of magnitude
higher than with columns 6, 7.
Comparison of Q packed bed chromatography and Q Membrane Adsorber6
Q Resin Q Membrane Adsorber
Flow rate 100–150 cm/h 450–600 cm/h
Protein loading (flow through) 50–100 g/l >3,000 g/m2 or
>10.9 kg/l
Buffer used 100% 5%
Cleaning validation Yes No
6 J.X. Zhou, T. Tressel (Amgen Inc.), Membrane Chromatography as a Robust Purification System for Large-ScaleAntibody Production, BioProcess Int. 09, 2005, p. 32–37.
7 H. Knudsen et al. Membrane Ion Exchange Chromatography for Process-Scale Antibody Purification, J. Chromatography A 907, 2001, 145–154
Virus clearance
This study5 also demonstrates excellent virus log reduction values with Sartobind Q.
This enables membrane adsorber technology to replace a Q column without compromising
critical viral clearance capabilities.
Murine Pseudorabies Reo Virus Minute Virus
Leukemia Virus type 3 of Mice
Virus (MuLV) (PRV) (Reo-3) (MVM)
50 ml virus > 5.57 ± 0.25 > 5.67 ± 0.17 > 7.28 ± 0.30 > 6.77 ± 0.24
spiked protein
sample
500 ml virus 6.29 ± 0.25 > 5.67 ± 0.23 > 7.53 ± 0.29 4.41 ± 0.37
spiked protein|
buffer solution
5 R. Zhang (Abgenix Inc.) et al, Viral Clearance Feasibility Study with Sartobind Q Membrane Adsorber for HumanAntibody Purification, BioProduction 2004, Munich Germany 26–27 October 2004.
Fig. 5: No size exclusion effects for viruses
Fig. 6: Oversized column compared to Sartobind capsule for polishing application
Capsules vs. Columns
6
Dimensions and connections
Capsule with 4 mm pico nano mini 5” 10”, 20”, 30” mega
bed height
Connector Luer Lock Luer Lock Sanitary 3’’ Sanitary 11’’ Sanitary 11’’ Sanitary 1’’
Capsule 31+ 11 mm 37+ 33 mm 87+45 mm 270+70 mm 365+100 mm (10”) 910+190 mm
(height+diameter) 620+100 mm (20”)
870+100 mm (30”)
Inlet connector female female 25 mm outer d 50.5 mm outer d 50.5 mm outer d 50.5 mm outer d10 mm inner d 15 mm inner d 36 mm inner d 36 mm inner d
Outlet connector as above as above as above 50.5 mm outer d as above as above
19 mm inner d
Hose barb version
Capsule (height+width) n. a. n. a. 103+45 mm 190+70 mm n. a. n. a.
Hose barb connectors n. a. n. a. 6–12 mm 14.5 mm n. a. n. a.
Recommended internal n. a. n. a. 6–10 mm 12 mm n. a. n. a.
diameter of flexible tube
n.a. = not availableRelated products are Sartobind MA units for laboratory use and the Sartobind System Jumbo line with 8 mm bed height (Sartobind nano 3 ml, 150 ml and Jumbo 5 l) with void volume optimization for bind & elute and polishing.
Technical data
Membrane
Base material Stabilized reinforced cellulose
Membrane thickness 275 μm
Pore size (nominal) >3 μm
Ion exchanger types – strong anion Q (quaternary ammonium)
R-CH2-N+(CH3)3
– strong cation S (sulfonic acid) R-CH2-SO3-
– anion exchanger (primary amine) -NH2
Capsule materials
Outer cage, inner core, Polypropylene
end caps, capsule housing,
nonwovens
O-ring in vent valve* Silicone
Bed height 4 mm
* only with 10”, 20”, 30” and mega capsules
Operation
Depyrogenation 1 N NaOH 30–60 min at 20°C
Autoclaving 121°C for 30 minutes for one cycle only
Maximum operating pressure 4 bar, 0.4 MPa, 58 psi
5 bar, 0.5 MPa, 72.5 psi (Sartobind STIC PA 5")
Fig. 7: Sartobind pico 0.08 ml for processdevelopment and virus studies
Fig. 8: Sartobind Q SingleSep nano 1 ml
Ordering Information
Sartobind Q capsules
Order no. Chemistry Description Quantity Bed Bed Nominal Frontal Protein Recom- Approx.
adapter inlet volume height membrane surface* binding mended Capsule
and outlet [ml] [mm] area area capacity** flow rate weight
[cm2] [g] [l/min]
Sartobind pico
92IEXQ42DD-11--D S Luer female 10 0.08 4 2.9 cm2 0.19 0.002 0.002 1.5
Sartobind nano
92IEXQ42DN-11 Q Luer female 1 1 4 36 cm2 2.4 0.029 0.03 10 g
92IEXQ42DN-11--A Q Luer female 4 1 4 36 cm2 2.4 0.029 0.03 10 g
Sartobind mini
92IEXQ42D4-OO--A Q hose barb 4 7 4 250 cm2 20 0.2 0.2 30 g
92IEXQ42D4-SS--A Q sanitary flange 4 7 4 250 cm2 20 0.2 0.2 30 g
Sartobind 5 inch
92IEXQ42D9-OO--A Q hose barb 4 70 4 2,500 cm2 160 2 1.9 220 g
92IEXQ42D9-SS--A Q sanitary flange 4 70 4 2,500 cm2 160 2 1.9 230 g
Sartobind 10 inch
92IEXQ42D1-SS Q sanitary flange 1 180 4 6,600 cm2 450 5.3 5.0 800 g
Sartobind 20 inch
92IEXQ42D2-SS Q sanitary flange 1 360 4 1.3 m2 900 10.5 10 1.4 kg
Sartobind 30 inch
92IEXQ42D3-SS Q sanitary flange 1 540 4 2 m2 1,350 15.8 15 2 kg
Sartobind mega
92IEXQ42DC3SS Q sanitary flange 1 1620 4 6 m2 4,050 48 50 5.6 kg
* Frontal surface area = average cylinder surface
** Typical dynamic binding capacity at 10% breakthrough was measured with BSA for Q | PA and lysozyme for S chemistry.
For Sartobind STIC PA 150 mM NaCl was added to the buffer.
7
8
Sartobind STIC capsules
Order no. Chemistry Description Quantity Bed Bed Nominal Frontal Protein Recom- Approx.
adapter inlet volume height membrane surface* binding mended Capsule
and outlet [ml] [mm] area area capacity** flow rate weight
[cm2] [g] [l/min]
Sartobind pico
92STPA42DD-11--D STIC PA Luer female 10 0.08 4 2.9 cm2 0.19 0.004 0.002 1.5 g
Sartobind nano
92STPA42DN-11--A STIC PA Luer female 4 1 4 36 cm2 2.4 0.050 0.03 10 g
Sartobind 5 inch
92STPA42D9-FF--A STIC PA sanitary flange 4 70 4 2500 cm2 160 3.5 1.9 230 g
Sartobind 10 inch
92STPA42D1-SS STIC PA sanitary flange 1 180 4 6600 cm2 450 9 5 800 g
Sartobind 30 inch
92STPA42D3-SS STIC PA sanitary flange 1 540 4 2 m2 1,350 27 15 2 kg
Sartobind mega
92STPA42DC3SS STIC PA sanitary flange 1 1620 4 6 m2 4,050 81 50 5,6 kg
Sartobind S capsules
Sartobind pico
92IEXS42DD-11--D S Luer female 10 0.08 4 2.9 cmÇ 0.19 0.002 0.002 1.5
Sartobind nano
92IEXS42DN-11 S Luer female 1 1 4 36 cm2 2.4 0.025 0.03 10 g
92IEXS42DN-11--A S Luer female 4 1 4 36 cm2 2.4 0.025 0.03 10 g
Sartobind mini
92IEXS42D4-OO--A S hose barb 4 7 4 250 cm2 20 0.175 0.2 30 g
92IEXS42D4-SS--A S sanitary flange 4 7 4 250 cm2 20 0.175 0.2 30 g
Sartobind 5 inch
92IEXS42D9-OO--A S hose barb 4 70 4 2,500 cm2 160 1.75 1.9 220 g
92IEXS42D9-SS--A S sanitary flange 4 70 4 2,500 cm2 160 1.75 1.9 230 g
Sartobind 10 inch
92IEXS42D1-SS S sanitary flange 1 180 4 6,600 cm2 450 4.6 5.0 800 g
Sartobind 30 inch
92IEXS42D3-SS S sanitary flange 1 540 4 2 m2 1,350 15.8 15 2 kg
* Frontal surface area = average cylinder surface
** Typical dynamic binding capacity at 10% breakthrough was measured with BSA for Q | PA and lysozyme for S chemistry.
For Sartobind STIC PA 150 mM NaCl was added to the buffer.
9
Accessories
Order no. Description
5ZGI--0001 Stainless steel holder for one 10, 20 or 30 inch capsule, 3 legs
5ZGLG-0004 Stainless steel holder for three 10, 20 or 30 inch capsules, 3 legs
5ZALB-0002 Stainless steel distribution adapter for three 10, 20 or 30 inch capsules
9ZAIAM0001 Stainless steel legs for mega, 3 items
1ZAOGV0003 Adapter Sanitary 25 mm 1–3“ to UNF10–32 to connect mini to LC systems, 2 items
1ZA---0004 Adapter Luer male to UNF – 10–32 female, PEEK, 1 item
Fig. 10: A Sartobind mega capsuleon stainless steel legs
Fig. 9: Adapter to connect mini with sanitaryconnection to liquid chromatography systems
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Ver
. 12 |
2011
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