CESI 8000 PLUS HIGH PERFORMANCE SEPARATION – ESI MODULE• Increased coverage fromlimited samples• High-resolution separationof polar analytes andcritical PTMs
• In-depth characterizationof proteoforms andprotein complexes

See What You’ve
Been Missing
CESI 8000 PLUS HIGH PERFORMANCE SEPARATION – ESI MODULE

For Research Use Only. Not for use in diagnostic procedures.

Expand theReach ofYour MassSpectrometer

The CESI 8000 Plus High Performance
Separations-ESI Module
The CESI 8000 Plus is a valuable accessory for your mass spectrometer whenanalyzing charged and polar compounds in every analytical laboratory:• Consumes minute samples with extraordinary sensitivity• Excels at charged and polar metabolites, peptides, and more
• Enables high-resolution separation of proteins and protein-complexes• Eliminates ion suppression bias so you can see what you’ve been missing• Robust and easy-to-use

CESI-MS “showed better separation efficiency andresolution with 100-fold less sample consumption
compared to an LC-based intact protein separation.”John Yates III
The Scripps Research Institute, La Jolla, CA, USA

CESI-MS “shows a separation efficiency forhigh-molecular-mass compounds like larger
peptides and proteins that is remarkably higher
than that seen with any LC method”“...high resolution CZE separation of intact glycoprotein
species coupled to MS has significant potential forthe in-depth characterization and quantitative analysisof biopharmaceutical proteoforms.”
Barry Karger
Barnett Institute, Northeastern University, Boston, MA, USA“CESI-MS is a high-end tool for Metabolomics
work in my lab”

“With only a 200 fmol injection, CESI-MS/MS
enables high quality mass spectra while completely
characterizing the primary structure of biotherapeuticproteins in terms of amino acid sequence, glycosylationand post-translational modifications. This allows
you to see more with less, and strengthens thepower of the methodology in the context ofbiosimilarity assessment.”

Dr. Rawi Ramautar
Leiden Academic Center for Drug Research, The Netherlands

Yannis-Nicolas Francois
University of Strasbourg, France

Herbert Lindner
Innsbruck Medical University, Austria

Connecting the CESI 8000 Plus to your MS is aseasy as plug and spray. Simply click the OptiMS
cartridge tip into the source adapter.

“With monoclonal antibodies having become sucha significant class of drugs, not just in the BioPharma
space but the entire Pharma, capillary electrophoresis
has become an indispensable tool for monitoringproduct related impurities and product variants of theseblockbuster drugs. Advances like the CESI 8000 now
allow for the coupling of CE and MS and provide
a solution for the desperately needed identificationof peaks separated by capillary electrophoresis.”
Timothy Blanc
Eli LillyMore Data from Less Sample
CESI-MS is particularly valuable for precious samples such as:
subcellular fractions, microdialysates, CSF, murine samples,CTCs, needle biopsies, FFPE archives, or highly toxic samples
such as ADCs and venoms.
• Unique multi-segment injection increases throughput 10X• Extraordinary sensitivity and high resolution enablemaximum information with minimal sample consumption(~50 nL per injection)

Multiple experimentsin the time of one, with
<1 µL sample injected.

Ion Intensity (%)

1002500

The unique multi-segment injection(MSI) feature significantly increases
throughput. Multiple injections
are separated by 60 seconds ofbackground electrolyte injections
for sequential MS analysis. This
technique is commonly used with
MRM targeted analyte detection.

Concentration Values in µM

50
1500

625
3120

9.50

10.50

156 78

11.50

39 19

12.50

Migration Time (min)Data from multiple samples in single MS detection run.

13.50

14.50

Each dot is related to a single MS/MS
fragmentation spectrum.

Example of high sensitivity
from a limited sample

CESI-MS/MS
1250

m/z of the peptides (Da)Using only 100 ng of yeast mitochondrial
extract, CESI-MS significantly increased peptide
identification, resulting in a substantial increasein the number and variety of proteins identified.

1100
950
800
650
500
350
30405060708090

CE migration time (min)Number of Proteins Identified

NanoLC-MS/MS

Number of Peptides Identified

49222

78412

5641201m/z of the peptides (Da)

1050
950
850
750
650
550
450

•

nanoLC-MS/MS•

CESI-MS/MS

Kindly provided by Yannis-Nicolas Francois, University of Strasbourg, France.

•nanoLC-MS/MS•

CESI-MS/MS

121620

24

28

3236

LC elution time (min)4044

Separate Charged / Polar Molecules
and Proteins in High Resolution

Measurements of highly charged
analytes pose considerable challenges
to current technologies. CESI is themethod of choice for high-resolution
separation of polar metabolites,
peptides and proteins rich in particularpost-translational modifications(such as glycosylation, citrullination,

CESI-MS for Anionic Metabolic ProfilingElectrophoretic separation combined with highly
efficient ESI supports a variety of applications,from investigating the anionic and cationicmetabolome, to the study of proteoforms,
glycoforms, and multimeric protein complexes.

O
62500023000

Ion Intensity

methylation, and phosphorylation).

CH2OH

21000

H19000HO17000

H
O

H
OH

H

H

OH

H

HOO1OPO

O

PO

OO
H

H
OH

H

H

OH

OH

D-Glucose-6-phosphate;pKa (strongest acidic): 1.22

O

D-Glucose-1-phosphate;pKa (strongest acidic): 1.16

15000

CH2

The separation of sugar phosphates in central
metabolism and baseline separation of isobaric
positional isomers such as citrate and isocitratedemonstrates CESI-MS’ high resolving power.
CESI is also the preferred technology for chiralcompound separations in a cost effective
manner compared to chiral LC columns.

O

13000

H2C

11000

O

PO

OO

9000
H7000
5000

CH2OH

HHOOH

HOH

D-Fructose-6-phosphate;pKa (strongest acidic): 1.49

3000
1000

12.5

13.0

13.5

14.0

14.5

15.0

15.5

16.0

16.5

17.0

17.5

18.0

Time (Min.)
Kindly provided by Dr. Rawi Ramautar, Leiden Academic Center for Drug Research,
The Netherlands.

Total Ion Electropherogram

3E+80

Peptides Identified

For complex peptide mixture
analysis, CESI excels at various
post-translational modifications(PTMs) revealing importantbiological functional variations.

Ion Intensity

6E+8113100156161■ Non-modified Peptides (n=688)

134■ Phosphorylated Peptides (n=57)

85■ Acetylated Peptides (n=688)

32

91013119764

32

1

11020

130

40

7642

15060

Time (Min.)
Kindly provided by Herbert Lindner, Innsbruck Medical University, Austria.

Reconstructed Intensity

Intact MAb Separation of IgG1 by CESI-MS
3000
2500
2000
1500
1000
500
0

1460e51465e51470e5Mass. Da

Reconstructed Intensity

3000

Deamidations

146389.09

2500
146569.16

2000
1500

0

Deamidations
146701.58

146156.71

Deamidations
and oxidation

146862.70
146981.51

1460e51465e5Mass. Da

2000

146573.04

1500146536.36

1000
500

146387.58

2500

1470e5146535.621000
146719.51

5000146189.93

1460e5146948.99

1465e51470e51475e5Mass. Da

Different glycosylation and modification profiles
For deep characterization of proteoforms, glycoforms, charge states, and PTMs, CESI is the only front end technologythat enables liquid phase high resolution separation of proteins, metal binding, and protein complexes. CESI-MS is
an invaluable tool to drive biopharmaceutical development and biochemical mechanistic studies.

Principles and DesignCESI

Schematic of CESI sprayer
ESI metal needle

The integration of capillary electrophoresis (CE) withelectrospray ionization (ESI) into a single dynamic
process within the same device.
CESI was developed as a new front-end separationand ionization technique, combining the highefficiency and ultra-low flow of CE with ESI. This ESI
optimization has shown to greatly improve assay
sensitivity while reducing ion suppression bias.1, 2

4.0E+18+–
HV

Porous tip
Static conductive liquid

The CESI 8000 Plus was developed in collaboration with mass spectrometry
researchers covering a number of applications. They sought to expand theirrange of detection and increase sensitivity. In order to achieve this, ultra-low
flow separation was integrated with mass spectrometry using proprietary
CESI-MS technology.
The sprayer, located within a protective housing of the OptiMS cartridge,
combines an intrinsically low-flow CE separation with electrospray ionization(ESI) within a single device.

3.5E+18

Sensitivity (counts/mol)

Capillary

3.0E+18
2.5E+18• The CESI OptiMS capillary inlet and outlet are of the same inner diameter

2.0E+18
1.5E+18• The electrical contact for CE separation as well as for ESI generationis achieved through the ESI metal needle filled with conductive fluid

1.0E+18
5.0E+17
0.0E+00~10 nL/min ultra low flow

~50 nL/min low flow

~150 nL/min medium flow~250 nL/min moderate flow~350 nL/min high flow

• Ultra low flow at the porous capillary tip instantly generates a fine
ESI spray when a low voltage ~ 1.25 KV is applied

Infused sample, Angiotensin I at 2 ng/mL in 10% acetic acid. ESI voltage, -1,350 V; detection range, 50-3000 m/z• The low voltage of CESI-MS reduces oxidation artifacts
1. Kelly RT, Page JS, Zhao R, Qian W, Mottaz HM, Tang K, Smith RD, Anal.Chem. 2008, 80, 143-149
2. Schmidt A, Karas M, Dulks T, J., Am. Soc. Mass Spectrom. 2003, 14, 492-500

CESI 8000 Plus mobile design
The CESI 8000 Plus comes equipped with anelectric height adjustable mobile lab bench.
Sprayer Tip

The OptiMS cartridge locks into place in the CESI 8000
Plus, and the tip simply clicks into the source adapter,creating an easy-to-use plug-and-spray for supported
SCIEX mass spectrometers.

SampleInlet

Service, support, and consumables
Our customer support organization has access tothe latest product updates, software revisions,methods and repair procedures. The following isavailable to further optimize your use of CESI-MS:• Application Training• Operation Qualification (OQ)
• Chemistries for MS as well as optical detection

Zero dead-volume design
A single continuous separation capillary is contained
within the OptiMS cartridge, with the same internal
diameter from sample to spray. This design:• Does not require lengthy column equilibration• Eliminates sample carry-over• Reduces sample-to-sample injection timeMultiple experiments / sample
analyses from as little as 5 µL

Multi-MS connectivity andmulti-detection capability

The nanoVials have been validated foruse with 5 μL of sample. With careful
pipetting, injections from as little as
1 µL are possible. CESI-MS has ultra-low
sample consumption, leaving mostof the material available for analysis
by orthogonal techniques.

In CESI-MS mode, the CESI 8000 Plus can beconnected to industry-leading SCIEX mass
spectrometers. In stand-alone CE mode, UV/VIS,PDA, or LIF detection can be selected. This allows
for the analysis of biologics using our CE-SDS,
cIEF, and glycan chemistries.

• Increases robustness

CE Assays and Method Development

LIF

UV/VisPDA

MS Analysis

 TripleTOF® 5600
& 6600 Series

 QTRAP®/Triple Quad 5500
& 6500 Series

CESI 8000 Plus Easy-to-Use Software

Ready for Use Within Minutes
Simply select one of the preloaded methods, follow visuals
to place samples and reagents,and hit START.

Visual and Intuitive
The layout of the user interface
mimics the instrument to enabledirect tracking of sample contents.

Flexible yet Expandable
Select programmed control
or direct control to maximize
throughput and optimize
separation performance.

Module Specifications
Dimensions:
Height: 29.2 in (74.2 cm)
Door Open: 38.8 in (98.6 cm)
Width: 25 in (63.5 cm)
Depth: 28.4 in (72.1 cm)
Height Adjustable Portable Lab
Bench with Memory Settings:
36 in x 30 in (91.4 cm x 76.2 cm)
Height adjustable from 27 in to 44 in(69 - 112 cm)
Weight (uncrated):
188 lbs (85.3 kg)
Electrical Requirements:
Voltage: 100 - 240 V; 50/60 Hz
Voltage Range:
1 to 30 kV programmable
at 0.1 kV increments
Current Range:
3 to 300 μA programmableat 0.1 μA increments
Operating Environment Range:
15 - 30° C
20 - 60% relative humidity(non-condensing)
Sample Trays:

Ordering Information
Sample Temperature
Adjustment Range:
4 - 60° C
Capillary Temperature
Adjustment Range:
15 - 30° C
Stand-alone Capillary Temperature
Adjustment Range:
15 - 60° C
Pressure Delivery Range:-5 to +100 psi
Minimum Required Sample
Volume:50 µL when using microvials
5 µL when using nanoVials

A98089: MS CESI-MS base unit
Includes CE separation module, OptiMS Sprayer technology, andUV detector ready for stand-alone or CESI-MS analysis. Also includes
computer, monitor, system controller pre-loaded with the CESI
8000 Plus software, electric height adjustable mobile lab bench
making switching between CESI-MS and other front end separation
instrument is swift and simple.
A98089 MD 2: CESI-MS base unit with UV and Diode Array
Detectors (for standalone CE applications)
A98089 MD 3: CESI-MS base unit with UV and LIF Detectors(for standalone CE applications)
A98089 MD 4: CESI-MS base unit with UV, Diode Array andLIF Detectors (for standalone CE applications)
B07367: Silica Surface OptiMS Cartridge
30 μm ID, 150 μm OD x 90 cm
Contains bare fused-silica capillary for high-resolution separationof peptides, glycans, amino acids and nucleotides/nucleosides
B07368: Neutral OptiMS Cartridge
30 μm ID, 150 μm OD x 90 cm
Contains covalently attached neutral hydrophilic capillary surface
enabling high resolution separation of intact proteins and protein/
peptide/small molecule/metal complexes.
5043467: nanoVial
5 µL starting volume validated, 1 µL starting volume demonstrated
Biologic Characterization Kits390953: SDS-MW Assay Kit/Purity/Sizing
477600: Carbohydrate Labeling and Analysis Assay
A58481: cIEF Peptide Marker Kit (pl Marker Kit)
A80976: Advancied cIEF Starter Kit
CESI OptiMS Adapter Kits
B07363: OptiMS Adapter for SCIEX Nanospray III source

2 x 48 CESI vials / 0.3 mL microvials
Buffer Tray:2 x 36 CESI vials

For more product information on our capillary electrophoresis
systems visit sciex.com/ceproducts.
For information on CESI 8000 Plus and supported MS models/sources,
contact your SCIEX representative.

Your Success is Our Success
We take it personally
As a SCIEX customer you have access to a world-class customer support organization.
Wherever you are, we’re there with you as a trusted partner to answer questions, provide
solutions, and maximize lab productivity.
Our customer support organization has access to the latest product updates, software
revisions, methods and repair procedures to make sure that you stay on top of your game.
AB Sciex is doing business as SCIEX.

When you have questions, we have answers.
Learn more at sciex.com/customersupport, or locate your local accountrepresentative at sciex.com/contactus© 2018 AB Sciex. For Research Use Only. Not for use in diagnostic
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RUO-MKT-03-7454-A 01/2018

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