agilentsolutionsforbiologiccharacterization · 1444.87 2890.8 1 g0f/g0f x10 4 0 1 2 3 4 5 6 6.5...

Our measure is your success.

High-productivity

workflowsolutions

for biopharmaceutical analysis

Agilent Solutions for Biologic Characterization

products | applications | software | services

2

Agilent Solutions for Biopharmaceutical Analysis

The tools and technologies you need to bringyour biologic to market.

As the leading provider of analytical solutions to

the biopharmaceutical industry, Agilent is helping you

provide safe, high-quality, high-efficacy therapeutic

products. And we’re helping you get them to market

faster and more cost-effectively.

We offer an extensive portfolio of end-to-end,

workflow-driven solutions across key process

development, scale-up, manufacturing and

QA/QC applications, including:

• Physiochemical characterization

- Intact NBE analysis

- Glycosylation and phosphoprotein analysis

- Amino acid analysis

- Peptide mapping

• Product-related impurity analysis

• Process-related contamination testing

• Protein and antibody QA/QC

• Stability and dosage form testing

3To learn more about Agilent Solutions for Biopharmaceutical Analysis, visit www.agilent.com/lifesciences/biopharma

We understand the complexity and the depth of the technical

challenges you’re facing. We address concerns with protein

aggregates and fragmentation, and we provide robust analytical

solutions for host cell DNA, viruses, leachables, extractables, cell

culture media and components, as well as the purified reagents

and chemicals needed to ensure highest confidence in your

results. Agilent’s broad technology portfolio includes:

• Liquid chromatography

• LC/MS

• Capillary electrophoresis

• Ion analysis (CE and HPLC)

• Isoelectric focusing

• Evaporative light scattering

• Gas chromatography

• GC/MS

• qPCR

• LC column technology (reversed phase, ion exchangeand size exclusion)

• Protein standards and protein processing kits

• Lab-on-a-Chip

Across the entire biopharmaceutical pipeline, Agilent-reliable,integrated solutions deliver superior sensitivity, selectivity, speedand ease of use and real world performance that can measurablyimprove your company’s productivity.

Services and software that support theregulated labWith a comprehensive compliance portfolio that includesIQ and OQ for a single instrument, multiple labs or global,multi-vendor implementations, Agilent is ready to helpyou optimize, automate and streamline complianceprocedures. We can improve the consistency and accuracyof your validation initiatives, while freeing up staff time,minimizing risk and reducing costs.

Agilent software solutions also support GLP/GMP and21 CFR Part 11 compliance with features such as built-inaudit trails, multi-user login security, configurable userpermissions and electronic signatures.

4

Physiochemical Characterization | Purity Monitoring and Stability

CE • CE/MS • LC • LC/MS • GC • GC/MS • ICP-MS

From the most advanced R&D to routine QA/QC, Agilent offers a

broad range of solutions for determining integrity, purity and molecular

weight of proteins and oligonucleotides. Complementary separation

tools, like the OFFGEL System and Bio-monolith LC columns, enable

highest resolution and fastest separation of intact proteins. OFFGEL

can also be used to diagnose peaks found in CIEF (capillary isoelectric

focusing) separations.

Our 6200 and 6500 Series LC/MS systems deliver industry-leading

performance, reliability and value. Agilent CE and CE/MS systems

extend your analytical options. Our 1200 Series LC systems

with ELSD detector allow both size exclusion and light scattering

techniques to be applied to aggregate issues.

Size Exclusion separation of AlbuminMonomer, Dimer and Aggregate usingthe ZORBAX GF-250/450 columns. Particles are specially treated to reduceprotein sticking and to increase long column lifetimes.

Agilent’s 1200 Series LC provides exceptional precision, resolution, reproducibilityand spectral quality at all flow rates, over a wide range of mobile phases.

DeLeenheer, A.P. et al. (1991) J. Pharm Sci. 80: 11.

Reprinted with publisher's permission

1. Aggregate

2. Albumin dimer

3. Albumin

12

3

Capillary electrophoresis using Laser-Induced Fluorescence allows highsensitivity protein profiling. Monoclonal antibody stability was monitored atdifferent incubation times with mercaptoethanol. Data courtesy of Picometrics.

FITC

t = 10 min.

0 3 6 9 12 15

t = 5 min.

t = 1 min.

t = 0 min.

Fluo

resc

ence

RFU

Minutes


Physiochemical Characterization | Integrity &Molecular Weight Analysis

LC/MS • LC-ELSD • CE • CE/MS • 2100 Bioanalyzer

Convenient molecular weight determination on the Agilent 2100 Bioanalyzer allows rapid, one-step methodology for your antibodies

and other biologics.

PAGE-less protein and antibody QC testing. In one simple step, you get details on proteinsize, stability, purity and concentration. Using pre-developed methods with a convenientreagent kit, the easily validated system analyzes 10 samples in less than an hour.

0

5

10

15

20

25

15 20 25 30 35 40 45 50

1000

0

500

15 20 25 30 35 40

0.05 %

Time [sec]Fl

uore

scen

ce

Highly sensitive protein analysis. The High Sensitivity Protein 250 kit for theAgilent 2100 Bioanalyzer offers four orders of magnitude in linear dynamic rangeand automatically detects and quantifies protein impurities down to 0.05%.

The Agilent 2100 Bioanalyzer lets you trade messy gels for convenient, cost-effective LabChip assays—a convenient and cost-efficient solution for testingand monitoring protein and antibody products.

1444.87

2890.81

G0F/G0Fx10 4

0

1

2

3

4

5

6

6.5148812.81

145922.00

147367.94

146329.69

146816.21147719.65

Counts vs. Deconvoluted Mass (amu)

145500 146000 146500 147000 147500 148000 148500 149000 149500 150000 150500 151000 151500

5.5

4.5

3.5

2.5

1.5

0.5

162.16 hexose unit

x104

0

1

2

3

4

5

6

7

8

148812.81

148974.97148840.65148916.37

148765.43

Counts vs. Deconvoluted Mass (amu)148750 148800 148850 148900 148950

G0F/G0F

G0F/G1F

6

High-confidence antibody characterization.Agilent’s Ultra High Definition (UHD) mass spectrometersprovide clarity needed to fully characterize intact antibodies. In the example above, the differingglycoforms can be clearly seen. MassHunter BioConfirm software (in this example showing thedeconvoluted spectrum of an intact mAb) uses sophisticated algorithms for automated confirmationand characterization of recombinant proteins or synthetic peptides.

50

40

30

20

10

0

0 2 4 6 8 10 min

Heavy-chain forms:Deglycosylated andC-terminal lysine removed

Light chains:Differing by twoamino acids

IgG1 treated with:DTTPeptide-N-glycosidase FCarboxypeptidase B

High resolution separations of antibodies. The Agilent Poroshell 300SB-C8 columns enhancepeak shape and recovery through simplified interaction with the bonded phase. The Poroshell300 column provides clear separation between heavy and light antibody chains of a monoclonalantibody treated with DTT, peptide-N-glycosidase F and carboxypeptidase B.

Physiochemical Characterization | Identity

LC • LC/MS • CE • CE/MS • Peptide Mapping • Amino Acid Analysis

Agilent’s CE and CE/MS high resolution capability enables thorough, confident characterization of proteins, oligonucleotides and glycans.

Our LC and LC/MS solutions are also widely used for identity confirmation via peptide mapping and amino acid analysis.


PeptideMapping. Agilent’s end-to-end peptide mapping solution starts with the 1200 Series LC andincludes trypsin to digest the proteins, columns to separate the peptides, detection systems—UVand TOF-MS—and sophisticated software to elucidate and correlate with theoretical sequences.

Rapid PeptideMapping using the Agilent 1290 Infinity LC. For high throughput, UHPLC technologyin the 1290 Infinity LC system enables full LC/MS peptide maps in less than 1.5 minutes.

Amino Acid Analysis. Agilent offers a complete solution for HPLC amino acid analysis—includingcolumns and reagent kits.

1. ASP2. GLU3. ASN4. SER5. GLN6. HIS7. GLY8. THR9. ARG10. ALA11. TYR12. CY2

13. VAL14.MET15.NVA16. TRP17. PHE18. ILE19. LEU20. LYS21. HYP22. SAR23. PRO

8

The Agilent Bio-Monolith DEAE column provides a rapid and accuratesolution for monitoring fermentation-related impurities. Host genomic DNA(peak 2) was monitored throughout a phage proliferation. This analysisprovides the optimal time to stop the bioreactor before the DNA beginsto degrade (peak 3), which cannot be removed through purification.

2 4 6 8

mAU

0

10

20

30

40

50

60

Time [min]

Heparin

Contaminant A

Contaminant B

High-speed, high-resolution CE separation of heparin. Agilent'sextended light path capillaries provide a significant sensitivityadvantage for the detection of heparin-related impurities in thisrapid, robust QA method.

6 th Charge State

Full length productand related salts

N+1 familywith linker

M-20

N-4 no linker3’phos,(5th)N-2 no linker (6th)

rG cleavage with linker, (5th)N-1 familywith linker

M-94

N-1 no linker1559.9

1569.81659.5

1697.2

1709.5

1722.1

1756.1 1772.8

0

2000

4000

6000

Intens.

1575 1600 1625 1650 1675 1700 1725 1750 1775 m/z

Counts (%) vs. Deconvoluted Mass (amu)

Process Impurities

Oligonucleotide analysis. In this LC/MS analysis of a synthesized 30-mer RNA aptamer, an expanded view ofthe 6th charge state reveals a variety of process impurities.

Impurity Testing | Product-Related Impurities

LC • LC/MS • CE • CE/MS • GC • GC/MS • ICP-MS

With the industry’s most complete—and most reliable—portfolio of instruments, columns and software, we deliver the assurance

you need to safeguard the quality of your therapeutic products.

Avoiding protein aggregates and fragmentation is a particular concern—and can be challenging. Agilent can help with LC, LC/MS,

microfluidic electrophoresis devices and size exclusion HPLC systems to determine these product-related impurities.


4.00 8.00 12.00 16.00

100000

200000

300000

400000

500000

600000

700000

800000

900000

1000000

Time-->

Abundance

TIC: liner cryomilled inj#2.D

Cap Liner, Scan data

4.00 8.00 12.00 16.00

100002000030000400005000060000700008000090000

100000110000120000130000

Time-->

AbundanceTIC: liner cryomilled inj#2.D

Cap Liner, SIM data

40 45 50 55 60 65 70 75 80 85 90 95 0

5000

10000

15000

20000

25000

30000

35000

40000

45000

50000

55000

60000

65000

70000

75000

80000

m/z-->

Abundance

Scan 1733 (9.831 min): �liner cryomilled inj#2.D

94

66

7340 6350534744 79 8860 847670

The StratageneMx3005P QPCR system offers unmatched flexibilityand capability to support the widest range of real-time QPCRapplications and chemistries.

The Agilent 7000B Triple Quadrupole GC/MS delivers femtogram-level detection of target compounds—along with maximum day-after-day productivity.

GC/MS headspace analysis of leachables. The SIM and scan dataclearly show the presence of phenol in a linear polymer material. Com-pound confirmation was made by comparison with a hazardous chemi-cals library.

Impurity Testing | Host Cell DNA, Viruses

Stratagene Quantitative PCR

With the addition of Stratagene real-time PCR, Agilent is able to offer complete workflow solutions to address contamination of host

cell DNA and viruses—a critical step in ensuring the purity of your biological organisms expressing your product. Stratagene Brilliant II

QPCR reagents offer superior sensitivity of detection and accommodate rapid cycling for fast, reliable results.

Impurity Testing | Column/Vessel Leachables and Extractables

LC • GC • GC/MS

With a 40-year history of leadership, Agilent offers the industry’s most trusted GC and GC/MS solutions. Our systems are widely used

to detect and identify residual solvents in cleaning processes, as well as volatile leachables from packaging and other sources.

The Agilent 1200 Series LC continues to set the standard for analysis of non-volatile process- and packaging-related contaminants,

such as cell culture media components.

10

Integrated software solutions for R&D, scale-up,QA/QC—and the enterprise.From instrument control to enterprise content management, Agilent recognizes the critical role software plays in

bringing your company’s products to market. Our software and informatics team is working towards a single goal:

To help you reduce the time, effort and cost to get from raw data to final insight.

Agilent ChemStation software

• Industry-standard chromatography data system for

Agilent instrumentation

• Robust, scalable CDS ideally suited for biopharmaceutical

applications from early product development to

quality control

Agilent MassHunter Workstation software

• Perfectly complements the best-in-class performance

of Agilent’s mass spectrometry solutions

• Single, easy-to-use interface handles basic qualitative

and quantitative tasks for all Agilent MS platforms

• Seamlessly integrates with application modules such

as Mass Profiler Professional and MassHunter

BioConfirm software

MassHunter BioConfirm software

• Combines powerful data processing algorithms with

sophisticated sequence and database matching tools for

fast, accurate identification of intact proteins, peptides

and their variants

• Includes a Large Molecule Feature Extractor (LMFE)

specifically designed for analyzing complex mixtures

of intact proteins or large oligonucleotides


Mass Profiler Professional

• The only chemometrics software package designed

specifically for mass spectrometry data

• Lets you easily import, normalize, compare and display

microarray and MS data, including GC/MS, LC/MS,

CE/MS and ICP-MS

• Lets you analyze different experiment types in a

single project

Kalabie ELN lets you spend less time on paperwork

and more time on science.

• Scalable, integrated platform for cross-team collaboration

• Simplifies and accelerates the R&D process and streamlines

data management

• Provides a robust IP protection framework

Agilent OpenLAB Enterprise Content Manager (ECM)

• Brings all your lab’s information resources together in a

scalable Web-based environment

• Provides comprehensive search functions and fast,

easy accessibility

For more information

Learn more:

www.agilent.com/lifesciences/biopharma

Buy online:

www.agilent.com/chem/store

Find an Agilent customer center in your country:

www.agilent.com/chem/contactus

U.S. and Canada

1-800-227-9770

[email protected]

Europe

[email protected]

Asia Pacific

[email protected]

Research use only. Information, descriptions and specifications in this publication aresubject to change without notice. Agilent Technologies shall not be liable for errorscontained herein or for incidental or consequential damages in connection with thefurnishing, performance or use of this material.

© Agilent Technologies, Inc. 2009Printed in USA November 9, 20095990-4542EN

Agilent Services keep your labrunning at peak productivity.Agilent Advantage Service not only protects your investment

in Agilent instruments—it connects you with our global

network of experienced service professionals dedicated to

helping you get the highest levels of speed, resolution and

performance from every system in your laboratory. Count

on us to provide the services you need across all stages of

your instrument’s life cycle, from installation and upgrade

to maintenance, qualification and repair.

You can also trust Agilent to deliver proven instrument

qualification and calibration services in support of GMP

and GLP guidelines. In fact, Agilent has been ranked #1 in

compliance by independent survey since 1997. With over

100,000 successful qualification deliveries, industry-leading

quality development processes and over a decade of practical

compliance experience, Agilent Compliance Services helps

assure confidence in your analytical results.

The Agilent Value Promise—

10 years of guaranteed value.In addition to continually evolving products, we offer

something else unique to the industry—our 10-year value

guarantee. The Agilent Value Promise guarantees you at

least 10 years of instrument use from your date of purchase,

or we will credit you with the residual value of that system

toward an upgraded model. Not only does Agilent ensure

a safe purchase now, we help ensure your investment is as

valuable to you in the long run.

The Agilent Service Guarantee

Should your Agilent instrument

require service while covered by

an Agilent service agreement, we

guarantee repair, or we will replace your

instrument for free. No other manufacturer or service

provider offers this level of commitment to keeping your

laboratory running at maximum productivity.

agilentsolutionsforbiologiccharacterization · 1444.87 2890.8 1 g0f/g0f x10 4 0 1 2 3 4 5 6 6.5...

Documents