zipchip™ system for selected thermo scientific™ mass

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ZipChip™ System for Selected Thermo Scientific™ Mass Spectrometers-- for CE/ESI-MS Analyses

2

Content

Resources and Ordering Information

ZipChip System Introduction

ZipChip Separation Mechanism

Biopharmaceutical Application Examples

Metabolomics Application and System Test

3

• The ZipChip system uses integrated

microfluidic technology to prepare,

separate samples by capillary electrophoresis (CE), and then

electrospray (ESI) analytes directly into

a mass spectrometer (MS)

• It is composed of the ZipChip interface

and the microfluidic chip

• ZipChip Interface directly mounts onto

the front end of a mass spectrometer

• The CE separation and ESI occur on the microfluidic chip

• ZipChip system is compatible with a

broad range of biomatrices such as growth media, cell lysates, blood,

plasma, and urine

• The ZipChip system uses integrated

microfluidic technology to prepare,

separate samples by capillary electrophoresis (CE), and then

electrospray (ESI) analytes directly into

a mass spectrometer (MS)

• It is composed of the ZipChip interface

and the microfluidic chip

• ZipChip Interface directly mounts onto

the front end of a mass spectrometer

• The CE separation and ESI occur on the microfluidic chip

• ZipChip system is compatible with a

broad range of biomatrices such as growth media, cell lysates, blood,

plasma, and urine

What is the ZipChip System?

ZipChip HS ZipChip HR

Separation channel length (cm) 10 22

Flowrate (nL/min) 150 150

Maximum # of injections per chip 125 125

On Chip De-salting capability Yes Yes

Integrated ESI Emitter Yes Yes

EEPROMS (recognize chip type and track usage) Yes Yes

Recommended use Small molecules or simple sample mixture Big molecules or complex sample mixture

Typical analysis time Up to 3 min Up to 15 min

4

Why the ZipChip System?

• Capillary Electrophoresis (CE) has unique advantages for certain analyses where traditional LC has its limitation

Separation of charge variants of intact antibodies/proteins

Effective separation of monosaccharides, glycans, and glycopeptides in a MS compatible condition

Separation of very hydrophilic molecules Separation of amino acid isomers

Analyses of samples in high salt condition

• Other CE instruments that can be coupled with MS are bulky with cumbersome operations and take longer analysis time

• They are not specifically designed to be coupled with MS therefore requires additional interface

• There is a lack of convenient CE/ESI device that offers fast and good CE separation for mass spectrometry analysis

• ZipChip is the ONLY commercially available portable and integrated CE/ESI source designed for mass spectrometers

Fast CE separation • Nano Spray Sensitivity • HRAM Mass Spectrometry

5

Unique Values and Common Applications

• Intact mAb and ADC/biotherapeuticscharacterization

Native condition Denatured condition

• mAb subunit analyses

• Glycomics and glycoproteomics

• Peptide mapping

• Metabolomics

• The only commercially available integrated and portable CE/ESI interface for MS

• Offers extremely rapid CE separations, nano-spray level sensitivity, and HRAM mass spectrometry in one platform

• Requires minimal sample preparation with on-chip desalting capability

• Consumes only picograms to nanograms of sample per analysis

Common Applications (not a full list)Unique Values

6

Comprehensive Portfolio

ZipChip Interface ZipChip Autosampler ZipChip Assay KitsZipChips

• Compatible with all Thermo

ScientificTM Exactive, Q

Exactive Orbitrap MS, and

LTQ Orbitrap MS instruments

• Data collection, processing

and reporting through

Thermo ScientificTM

Xcalibur and BioPharma

Finder software

• Two versions: Autosampler

operation version and manual operation version

• Disposable chips

• Good for up to 125

injections

• Two types: HR chip and HS chip

• Each chip is a single piece

of glass similar as a

microscope slide, housed in PEEK polymer

• Microfluidic channel and

electrospray is integrated

into both types of chips• 5 chips per pack

• 3 types of pre-made assay kits

are designed for intact antibody,

peptides, and metabolites

analyses, respectively• Each box of assay kit contains

5 bottles of BGE and 5 bottles

of sample diluents.

• Individual bottles can be directly loaded into ZipChip

autosampler

• Each individual bottle contains

enough volume for 100 runs• Each box is good for 500 runs

• Required to have for

ZipChip Interface

autosampler version

• Fully automated and controlled by the ZipChip

software

• Conduct chip priming,

sample and BGE loading, and chip flushing

• Compatible with both 48-

vial plate and 96-well plate

DryDock

• A simply drying

accessary

• No electrical

component• Utilize the MS gas to

push air through the

ZipChip channels

removing residual fluid

• Dry 1 chip each time

7

ZipChip-MS Analysis Workflow

Place ZipChip and prepare the system

3

Simple Sample Prep

2

Set up sequence and collect CE-MS data

4

Select proper assay kit and ZipChip for your experiments

1

8

Content






9

Anatomy of ZipChip

MS Inlet

Glass microfluidic chip in PEEK housing

+HV1

+HV2

Well 1Waste

Well 2BGE

Well 3Sample

Well 4BGE

Well 2BGE

Well 3Sample

Well 1Waste

Well 4BGE

10

+HV1

+HV2

Well 1Waste

Well 2BGE

Well 3Sample

Well 4

BGE

Sample Injection

10 µM rhodamine-6G in BGE + 100 mM ammonium acetate

11

Sample Separation

• High voltage applied to Wells 2 and 4

• HV1 and HV2 determine field strength

• Field strength drives the ZipChipseparation

+HV1

+HV2

Well 1Waste

Well 2BGE

Well 3Sample

Well 4BGE

��

6��

q - chargeη - viscosity - hydrodynamic radius

�� !" #$% &'$ℎ �)*1 , )*2

.ℎ&& ! / &'$ℎ

For ZipChip analysis analytes must be positively charged in solution

Cations: Migrate to ESI emitter and are electrosprayed

Anions: Migrate to Well 2

Neutrals: Do not migrate. Pushed to waste by EOF

--

++

Residual EOF++

++++++

+HV1

ESI Emitter

Well 1

(Waste)

Well 3

(Sample)

Well 2 (BGE)

--

------

(Electro-osmotic Flow)

12

Content






13

The ZipChip System with Thermo ScientificTM Q ExactiveTM BioPharma Platform

BioPharma Option is available for Q Exactive Plus, Q Exactive HF and Q Exactive HF-X mass spectrometers

14

The ZipChip system coupled with the

Q Exactive BioPharma platform can

quickly analyze intact mAbs in native, partially denatured, and fully

denatured conditions to support

biotherapeutics characterizations

under a diverse range of conditions

• CE/ESI-MS analysis can be

completed within 3 minutes

• High resolution accurate mass spectra in intact native, partially

denatured, and fully denatured

states on the Q Exactive

Plus/HF/HF-X MS with BioPharmaoption are confidently achieved

• Sample consumption can be as low

as pico grams to nano grams

• Major glycoforms are identified by

BioPharma Finder software

G0/G0F

G0F/G0F

G0F/G1F

(G1F/G1F) or (G0F/G2F)

G1F/G2F

1.0 ppm

3.9 ppm

1.0 ppm

7.5 ppm

2.4 ppm

0

20

40

60

80

100 148056.78

148218.15

148379.38

147910.25 148542.25

147800 148200 148600Mass

Rela

tive

Inte

nsity

1 2 3

Time (min)

0

100

0

100

Rela

tive A

bundance

0

1000.96

2.35

2.51

a

b

c

Fully denatured

Partially denatured

z=26

z=30

z=49

2000 4000 6000 8000m/z0

20

40

60

80

1000

20

40

60

80

100

Re

lative

Ab

un

dan

ce

0

20

40

60

80

1005695.4

5923.2

5484.6

5288.7 6183.7

3022.6

3085.5

3151.2

3444.2

4936.2

4777.1

5106.4

4627.83897.2 5288.8

3797.4

Native

5660 5700 5740m/z

5695.4

5701.7

5707.7

5689.8 5714.0

a

b

c

1 ng of sample

injected

160 pg of sample

injected

160 pg of sample

injected

Full MS spectra from Q Exactive HF-X MS with BioPharma Option

Electropherograms of intact Trastuzumab Deconvoluted spectrum

z=38

Intact mAb - Trastuzumab Analysis in HMR mode

MS data was acquired on a Q Exactive HF-X MS

with BioPharma OptionCE separation was achieved on ZipChip HR

with 10mM Ammonium Acetate and 10% Isopropanol (native),

Intact Antibody Assay kit (partially denatured), or Peptides Assay Kit (fully denatured)

Resolution: 30K

15

Electropherogram of intact infliximab:

0 2 4 6 8Time (min)0

20

40

60

80

100

Rela

tive A

bundance

2.6

4.53.5

0 Lys2 Lys1 Lys

5700 5720 5740m/z

0

50

1000

50

100

Rela

tive A

bundance

0

50

1005722.8

5717.9

5713.0

zoom

z=26Full MS spectra from Q Exactive HF-X MS with BioPharma Option

5000 6000m/z

0

50

1000

50

100

Rela

tive A

bundance

0

50

1005722.85510.8

5314.1 5951.7

5717.95506.2 5946.6

5309.5

5713.05941.45501.4

5305.0

z=26

z=25

z=27

z=28 RT 2.6 min

RT 3.5 min

RT 4.5 min

2 Lys

0 Lys

1 Lys

0

20

40

60

80

100148767.8

148511.9148929.2

148639.7

148673.3148801.2

148835.7 149092.6148964.9

148564.0149031.6 149199.3148311.6 149364.3 149481.5

148300 148500 148700 148900 149100 149300 149500

Mass

0 2 4 6 8Time (min)

0

20

40

60

80

100

Rela

tive A

bundance Sliding window deconvolution

with RespectTM algorithm

Rela

tive I

nte

nsity

[G0F/G0F] 2 Lys

[G0F/G1F] 2 Lys

[(G0F/G2F)/(G1F)2] 2 Lys

-Lys

-Lys

Deconvoluted MS spectrum

Theoretical and experimental masses of infliximab

Glycoforms of lysine variants

Lys-Variant Glycoform Theor. average mass Exp. average mass ∆ Mass (ppm)

2 Lys G0F/G1F 148930.7 148929.2 9.8

2 Lys G0F/G0F 148768.5 148767.8 4.8

2 Lys (G1F)2 or (G0F/G2F) 149092.8 149092.6 1.3

1 Lys G0F/G0F 148640.3 148639.7 4.3

1 Lys G0F/G1F 148802.5 148801.2 8.6

1 Lys (G1F)2 or (G0F/G2F) 148964.6 148964.9 -1.8

0 Lys G0F/G0F 148512.2 148511.9 1.6

0 Lys G0F/G1F 148674.3 148673.3 6.8

0 Lys (G1F)2 or (G0F/G2F) 148836.5 148835.7 5.1

The ZipChip system coupled with the Q

Exactive MS BioPharma platform is

unique and powerful to separate and identify different intact antibody charge

variants in native and denatured

conditions

• Baseline separation of intact mAb

charge variants resulting from different

levels of Lys-clipping can be achieved

within three minutes by ZipChip system

• High resolution accurate mass spectra

of all lysine variants are confidently detected on Q Exactive Plus/HF/HF-X

MS with BioPharma option

• Three major glycoforms from each of the three lysine variants are identified

by BioPharma Finder with mass

accuracies better than 10 ppm

3 ng of sample injected

Intact mAb - Infliximab Analysis in HMR mode



with 10mM Ammonium Acetate in 10% Isopropanol as BGE

Resolution: 30K

16

The rapid separation and accurate

identification of highly differently

abundant charge variants can also be

consistently achieved by the ZipChipsystem and Q Exactive BioPharma

platform

• Near baseline separation of intact NIST mAb charge variants with abundance

ranging over 2 orders of magnitudes can

be achieved by the ZipChip system

• High resolution accurate mass data of

each Lysine variant is confidently

obtained on Q Exactive Plus/HF/HF-X MS

• Glycoform with abundance as low as

0.16% of the base peak can be detected and identified

• All 5 major glycoforms from each of the

three different Lysine variants are identified by BioPharma Finder

Intact NIST mAb Analysis in HMR Mode

4920 4940 4960 4980

m/z

0

20

40

60

80

100

0

20

40

60

80

100

Rela

tive A

bundance

0

20

40

60

80

1004941.03

4946.44

4935.68

4951.80

4957.22

4945.27

4950.62

4939.84

4956.07

4961.46

4949.53

4944.11

4960.26

4965.69

NIST mAb + 0 Lys

NIST mAb + 1 Lys

NIST mAb + 2 Lys

+30

2.0 3.0Time (min)

20

40

60

80

100

Rela

tive A

bunda

nce

2.55

2.52

2.49

0

0 Lys

1 Lys

2 Lys

4600 4800 5000 5200 5400

m/z

0

50

100

0

50

100

Rela

tive A

bundance

0

50

1004941.03

4781.69

5111.40

4632.30

4957.22

5293.89

4802.31

4945.27

5115.804785.75

5298.444636.194966.75 5138.04

4806.46

4949.535120.25

4789.87

5308.79

4640.21

4971.14

4810.97

5142.36

0.65nL of 0.1ug/ul

NIST mAb injected

Protein Name Modification Average Mass

(Da)

Theoretical Mass

(Da)

Matched Mass Error

(ppm)

Intensity Relative

Abundance

RT

(min)

NIST 1xG0F_G0F 148038.8 148037.1 11.5 1.28E+09 58.82

NIST 1xG0F_G1F 148200.0 148199.3 5.0 2.18E+09 100.00

NIST 1xG1F_G1F 148362.1 148361.2 6.0 1.82E+09 83.26

NIST 1xG1F_G2F 148522.7 148523.5 5.8 1.00E+09 46.00

NIST 1xG2F_G2F 148684.6 148685.7 7.2 4.84E+08 22.21

NIST_plus1K 1xG0F_G0F 148165.6 148165.3 1.9 7.66E+07 3.51










2.42

2.49

2.55

Similar MS data were obtained on both Q Exactive Plus MS and Q

Exactive HF MS equipped with BioPharma OptionCE separation was achieved on ZipChip HR

with Intact Antibody Assay Kit

65pg of sample

injected

Resolution: 17,500

17

0

50

100150931.7

151091.8152048.7

151888.7

151252.7149974.5 150134.3

153007.0152210.5

150294.9152845.5

153168.3149176.3149015.4

151314.3153964.5149336.8

154126.3150358.5 152272.1151411.8 152371.5 153802.9150453.7154921.9148220.6

150580.1 153328.4 155143.3151538.8149499.1 154150.3149559.6148379.0 154758.8 155308.0

148871.3

154185.4152497.4

154282.5

148440.6 151744.1 155805.4153467.9 154661.2

155718.9

155474.7

154922.8

148500 150000 151500 153000 154500 156000

2.0 3.0 4.0 5.00

50

1003.34

1.0

Heterogenetic ADCs can be

successfully characterized within 1

minute without sample pre-treatment

by the ZipChip system and Q Exactive BioPharma platform, and

the powerful BioPharma Finder

software

• Powerful sliding window capability

enabled by BioPharma Finder

software

• All of the different forms of

Trastuzumab Emtansine can be

analyzed in less than 40 seconds with only 3 nanograms of sample

injected

• No sample pre-treatment required

• The calculated average DAR value

is consistent with previous published

data

Antibody-Drug Conjugate (ADC) Analysis in HMR Mode

4600 4650 4700 4750 4800 4850 4900 4950 5000 5050 5100 5150 5200 5250 5300 5350 5400 5450 55000

50

1005037.42

4874.895005.444905.77

4844.05 5069.29

5042.76

4911.01

4967.86

5074.72 5178.01 5205.514722.68 5101.284752.56 4813.01 5106.594782.45

5244.085145.03

5249.654692.75 5133.15

5271.445277.07

4995.22

5362.97 5391.495282.564608.56 5397.234637.63 5328.825304.57

4657.80 5431.315436.84

5471.365499.94

Electropherogram of Trastuzumab Emtansine

Deconvoluted spectrum

m/z

Re

lative

Ab

un

dan

ce

DAR 0 DAR 1 DAR 2 DAR 3 DAR 4 DAR 5 DAR 6 DAR 7 DAR 8

G0F/G1F DAR Form Mass Accurancy (ppm) Relative abundance

DAR0 10.72 9.95

DAR1 7.45 33.47

DAR2 1.23 58.97

DAR3 2.37 79.90

DAR4 5.63 69.92

DAR5 9.08 49.14

DAR6 13.89 28.97

DAR7 9.93 12.64

DAR8 8.41 2.83

Average Drug-to-Antibody Ratio (DAR) is 3.47

MS data was acquired on a Q Exactive HF with BioPharma

Option

CE separation was achieved on ZipChip HR with Intact

Antibody Assay Kit

Resolution: 15KMS spectrum

18

Fast, sensitive, and accurate antibody

subunit analysis can be

accomplished by the ZipChip system

and Thermo Scientific Q ExactivePlus/HF/HF-X MS platform

• Separation of infliximab mAb

subunits can be achieved in 3 minutes by ZipChip system

• The sliding window method combined

with Thermo Scientific™ Xtract™ deconvolution algorithm in

BioPharma Finder software enables

monoisotopic mass determination of each subunit

• Lysine variants and their major

glycoforms of the subunits can be identified by BioPharma Finder

software

IdeS digested + reduced infliximabIdeS digested infliximab

Subunit Monoisotopic theo. mass Experimental mass ∆ Mass (ppm)

Light chain 23424.3946 23424.4671 3.1

Fd' 25631.5325 25631.5644 1.2

Fc/2 G0F 25316.5863 25316.6345 1.9

Fc/2 G1F 25478.6391 25478.5753 2.5

(Fc/2 G0F) -Lys 25188.49131 25188.5763 3.3

(Fc/2 G1F) -Lys 25350.54413 25350.5246 0.8

F(ab')2 98146.3 (av.) 98146.8 -4.8

Fc/2 G0F (2 SS bonds) 25312.5550 25312.5524 0.1

Fc/2 G1F (2 SS bonds) 25474.6078 25474.5138 3.7

(Fc/2 G0F) -Lys (2 SS bonds) 25184.4600 25184.4311 1.1

(Fc/2 G1F)- Lys (2 SS bonds) 25346.5128 25346.4730 1.6

2.0 2.2 Time (min)0

20

40

60

80

100

Rela

tive A

bundance

2.11

2.14

2.26

(Fc/2) -Lys

F(ab’)2

Fc/2

120 k15 k

z=44

z=45 z=42

z=43

2000 2500m/z

0

20

40

60

80

100

Rela

tive A

bundance

2283.5

2231.7

2182.0z=41

z=46

z=48

Deconvolutionwith Respect 98146.8

98000 100000Mass

0

25312.5524

25184.4311

25474.513825346.4730

25100 25300 25500Mass

20

40

60

80

100

Rela

tive I

nte

nsity

(Fc/2 G0F) -Lys

Fc/2 G0F

(Fc/2 G1F) -LysFc/2 G1F

F(ab’)2

2.5 3.0 3.5 Time (min)

0

20

40

60

80

100

Rela

tive A

bundance 3.42 3.48

3.06

LC

Fd’(Fc/2) -LysFc/2

120 k

800 900 1000 1100m/z

0

50

100

0

50

100

Rela

tive A

bundance

845.43905.71

975.301014.31

1102.43

901.17970.37

1009.23

1096.90

841.13 (Fc/2) -Lys

Fc/2

LC

Fd’

1000 1500m/z

0

50

100

0

50

100

Rela

tive A

bundance

885.41 1026.87

1069.61

1166.81

938.54

1066.35

1172.93

1303.14

869.10

23424.467125631.5644

20

40

60

80

100

Rela

tive I

nte

nsity

Mass 2500024000

Fd’LC

(Fc/2 G0F) -Lys

Fc/2 G0F

(Fc/2 G1F) -LysFc/2 G1F

0

1200 1400m/z

0

50

100

0

50

100

Rela

tive A

bundance

1327.21401.1

1261.0

1483.31201.0

(Fc/2) -Lys

1334.11267.4

1408.21207.0

1490.9

Fc/2

IdeS

dig

est

IdeS

dig

est

plu

s r

eduction

mAb Subunits Analysis in Protein mode



with Peptides Assay Kit

19

0

50

100

1 2 3 4 5 6 7 8 9 10 11

The combination of ZipChip sample

separation, Q Exactive Plus/HF/HF-X

MS platform produced HRAM MS and

MS/MS spectra, and BioPharma Finder software enables fast and accurate

peptide identification

• Plug and play ZipChip delivers stable nano spray and nano spray level

sensitivity

• CE-MS/MS analysis can be completed in 10 minutes

• Only a few nanograms of sample are sufficient for the analysis

• 98% sequence coverage based on

MS/MS data for the light chain and heavy chain is confidently achieved

ProteinsNumber of MS

Peaks

MS Peak

Area

Sequence

CoverageAbundance

NSIT mAb light chain 141 26.4% 100.0% 41.67%

NIST mAb heavy chain 339 60.5% 97.6% 56.35%

Unidentified 1441 12.6%

1.2 ng of sample injected

NIST mAb light chain

NIST mAb heavy chain

Unidentified

Peptide Mapping Analysis in Standard Mode

Re

lative

Ab

un

dan

ce

Time (Min)

Electropherogram of Digested NIST mAb

MS data was acquired on a Q Exactive Plus MS with

BioPharma Option

CE separation was achieved on ZipChip HR with Peptides

Assay kit

MS resolution: 60K

MS/MS resolution: 15k

20

Content






21

Complete 20 Amino Acids Analysis

Extremely rapid separation and

accurate identification of all 20 amino

acid mix can be confidently achieved by

the ZipChip system and Thermo Scientific Q Exactive

/Focus/Plus/HF/HF-X MS platform

• Separation of 20 amino acids can be completed in 2 minutes by the ZipChip

system

• Isomers, Leucine(L) and Isoleucine(I), can be baseline separated

• Co-elueted amino acids can be identified with HRAM mass

spectrometry

• Femto grams level of sample is sufficient for each analysis

MS data was acquired on a Q Exactive MS

CE separation was achieved on ZipChip HS with Metabolites

Assay kit

Electropherogram of 10uM Promega 20 AA Mix

1.0 1.2 1.4 1.6 1.8 2.0 2.2Time (min)

0

10

20

30

40

50

60

70

80

90

100

0

10

20

30

40

50

60

70

80

90

100

Rela

tive A

bu

nd

an

ce

0.95

0.991.63

1.351.60

1.68

1.82

1.201.87

Base Peak

20 AA Extraction

1.54

1.72

K

R

H

GA

V IL

S

M,T

0.90

N,P

F

W,QY

E,CD

120 125 130 135 140 145 150 155 160 165 170 175 180m/z0

50

100

Rela

tiv

e A

bu

nd

an

ce

150.05851

120.06568

M

T

140 145 150 155 160 165 170 175 180 185 190 195 200 205 210 215 220 225 230m/z

0

50

100205.09689

147.07660wQ

116 117 118 119 120 121 122 123 124 125 126 127 128 129 130 131 132 133 134m/z0

50

100116.07079

133.06086

N

P

122 124 126 128 130 132 134 136 138 140 142 144 146 148m/z0

50

100 148.06065

122.02702C

E

1.381.41

I L

22

Linear Dynamic Range Test of Arginine and Valine

y = 1E+08x + 2E+07R² = 0.998

0,E+00

2,E+08

4,E+08

6,E+08

8,E+08

1,E+09

1,E+09

0 2 4 6 8 10 12P

ea

k A

reaConcentration (μM)

Arginine (R) 10nM~10μM

y = 1E+08x + 4E+06R² = 0.9999

0,E+00

2,E+08

4,E+08

6,E+08

8,E+08

1,E+09

1,E+09

0 2 4 6 8 10 12

Pe

ak

Are

a

Concentration (μM)

Valine (V) 10nM~10μM

Minimum sample loading of 40

attomole Arginine and Valine can be

achieved by the ZipChip system and

Thermo Scientific Q Exactive/Focus/Plus/HF/HF-X MS platform

• Arginine and Valine calibration curves

provide good linearity over the range of 10 nM to 10 µM

• Lowest sample injection amount on

the linear curve is 40 attomoles (4nL of 10 nM sample injected)


CE separation was achieved on ZipChip HS with

Metabolites Assay kit

23

Quick Analysis of Amino Acids in Gibco™ DMEM Cell Culture Media

K

R

H

G

V

I

L

S

M, TF

0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1 2.2

Time (min)

0

20

40

60

80

100

Re

lative

Ab

un

dan

ce

0.83

147.11197

1.70147.075581.35

132.10205

0.64

157.08342 1.38132.102050.88

175.118701.32

118.08601

1.64

166.086001.55

120.066160.69

105.003790.92

156.07758

1.84

182.081851.44

106.05044 1.94

241.028920.75

104.107061.07

76.03958

NL: 4.49E8

Base Peak MS Q

YW1.67

205.09830

choline

The ZipChip system and Thermo

Scientific Q Exactive

/Focus/Plus/HF/HF-X MS platform can

rapidly separate and identify the amino acids presented in cell culture

media

• Sample can be directly analyzed after simple filtration and dilution

• On chip desalting capability allows

direct analysis of samples in high salt condition without pre-treatment

• Separation can be completed in 2 minutes by the ZipChip system

• All 15 amino acids presented in the

cell culture media are identified and results match up with the aa

composition indicated in the

commercial sample spec sheet


CE separation was achieved on ZipChip HS with

Metabolites Assay kit

C-C

24

Calibration Curve of Internal Standard in Cell Culture Media

y = 2E+07x + 484283R² = 0.9996

0,E+00

5,E+07

1,E+08

2,E+08

2,E+08

3,E+08

0 2 4 6 8 10 12

Sig

na

l In

ten

sit

y

Concentration (μM)

Concentration range: 50 nM to 10μM

Val-d8Accurate quantification of amino

acids in cell culture media with stable

isotope labeled internal standard can

be achieved by the ZipChip system and Thermo Scientific Q Exactive

/Focus/Plus/HF/HF-X platform

• Deuterated Valine standard, Valine-d8, was spiked into the crude cell

culture media

• High linearity calibration curve with concentration range from 50nM to

10µM can be established.

MS data was acquired on a Q Exactive Classic MS


Assay kit

25

Carryover Test by Monitoring Arginine

0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0

Time (min)

0

20

40

60

80

100

0

20

40

60

80

100

0

20

40

60

80

100

Rela

tive A

bundance

0

20

40

60

80

100

0.95175.119

0.99156.077 1.34

118.0861.60

116.0711.37

132.1021.53

150.058 1.64205.097 1.78

182.0811.2090.055

1.84134.045

1.43106.050

0.95175.119

0.96175.119

NL: 1.51E9

Base Peak MS

NL: 1.51E9Base Peak m/z=175.118-175.120 MS

NL: 1.43E8

Base Peak MS

NL: 8.21E5Base Peak m/z=175.118-175.120 MS

10µM AA mix

Extraction of Arg

Extraction of Arg

Blank

The ZipChip system has extremely

low carryover

• Blank injections interleaved between 10 µM AA mix injections

• Extracted ion electropherogram of

Arginine from sample injection and blank injection were used to calculate

the carryover

• Observed carryover of Arginine is only 0.054% or less

MS data was acquired on a Q Exactive Classic MS


Assay kit

26

Migration Time Reproducibility Test Using Arginine (R) Standard

0,5

0,7

0,9

1,1

1,3

1,5

1 11 21 31 41 51 61 71 81 91

Mig

rati

on

Tim

e (

min

)

Injections

1st to 50th injections from a single vial RSD= 1.76%

51st to 100th injections from 96-well plate RSD= 0.84%

RSD= 1.85% (1st to 100th injections)

RSD=0.54% (8th to 50th injections)

Operation of the ZipChip system

through ZipChip autosampler can

achieve good reproducibility

• The migration time of Arginine was

monitored throughout 100 runs; the

overall RSD value of the migration

time is 1.85%

• It takes 5-10 injections for the system

to get stabilized

• First 50 injections are from the same

sample vial; the RSD value of the

migration time is 1.76%; the RSD value dropped to 0.54% if excludes

the first 7 injections

• Last 50 injections are from 50 different sample wells; the RSD value

of the migration time is only 0.84%



Assay kit

27

Content






28

Consumable Spec sheet

Application Note

www.thermofisher.com/zipchip

Hardware Spec Sheets

CE/ESI-MS Brochure

Available Resources

908devices.com/products/zipchip

29

Ordering Information

Product P/N

Hardware

ZipChip Interface (Autosampler version) 00950-01-00492

ZipChip Interface (Manual version) 00950-01-00493

ZipChip Autosampler 00950-01-00494

DryDock 00950-01-00518

Consumables

Installation Test Kit 00950-01-00510

ZipChip HS 00950-01-00498

ZipChip HR 00950-01-00499

ZipChip Metabolites Assay Kit 00950-01-00500

ZipChip Peptides Assay Kit 00950-01-00501

ZipChip Intact Antibody Assay Kit 00950-01-00502

30

Thank you!

Any Questions?

zipchip™ system for selected thermo scientific™ mass

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