ACQUITY UPLC H-Class Bio

BIOCOMPATIBLE UPLC SYSTEM

PURPOSE BUILT FOR THE ANALYSIS OF PROTEINS, PEPTIDES, NUCLEIC ACIDS, AND GLYCANS

Increasing complexity of modern biotherapeutic characterization, monitoring, and quality control assays

require even more demanding analytical techniques to accurately define and assess critical quality

control attributes (CQAs) related to product safety, stability, and efficacy.

A biocompatible variant of the ACQUITY® UPLC® H-Class System, the ACQUITY UPLC H-Class Bio System

delivers the same high quality separations you have come to expect from UPLC Technology, with the

flexibility and ruggedness to run all the chromatography modes you require – ion exchange (IEX),

size exclusion (SEC), hydrophilic interaction (HILIC), or reversed phase – all on the same system.

www.waters.com/hclassbio

With its flow-through needle injector and robust, quaternary solvent delivery system, the ACQUITY UPLC H-Class Bio System has been specifically engineered with a bio-inert flow path and Auto•Blend Plus™ Technology to deliver the benefits of UPLC in a system optimized for the analysis of proteins, peptides, nucleic acids, and glycans. The complete flow path is engineered with corrosion-resistant materials, helping your lab extend the lifetime of your instrument and maintain optimal bioseparations performance.

Analytical laboratories around the world are transferring existing methods to the ACQUITY UPLC H-Class Bio System. Download the Waters Bioseparations Applications Notebook to learn more about our tools, techniques, and insights for biopharmaceutical analysis.

■■ Reversed phase for peptide mapping and protein separations

■■ SEC for detecting clips and aggregates

■■ IEX for charge variants

■■ HILIC for released glycan analysis and glycoprotein/glycopeptides separations

■■ HIC for ADC analysis

■■ Multidimensional liquid chromatography (2D-LC)

■■ Reversed phase for amino acid analysis

www.waters.com/hclassbio

Low dispersion

True UPLC performance with band spread of less than 10 µL for highest chromatographic resolution.

Detection optimized for UPLC separations

UPLC-optimized detectors to match your application needs, including photodiode array, UV/Vis, fluorescence, refractive index, evaporative light scattering, and mass detection. Ultra-low-dispersion detectors designed to maintain peak integrity.

Optional fraction collection

Temperature-controlled and compatible with narrow UPLC peaks.

Versatile column management

A variety of column heaters and multi-column managers (with column switching) offer the lowest possible dispersion and precise temperature management for the highest retention time precision. This ensures that you will produce accurate results using methods that can be seamlessly transferred to other laboratories – down the hall or around the world.

Bio-inert flow-through- needle injector

Volume range of 0.1 to 1000 µL and ultra-low sample carryover performance compatible with your most sensitive LC and LC-MS applications.

Auto•Blend Plus Technology

Continuous blending of four solvents in any combination or proportion. Eliminates the variability caused by manual preparation of solvents. Manages the desired pH and ionic strength requirements for the selected mobile phase.

Quaternary solvent blending

The bioQuaternary Solvent Manager’s bio-inert flow path accommodates aqueous, high ionic strength solvents. Delivers repeatable gradient separations at pressures up to 15,000 psi. The addition of the optional, six-port solvent valve greatly expands solvent choices allowing for optimal method conditions.

Flexible sample support

Use either vials or ANSI well plate formats. Maximal sample capacity of 768 in two 384-well plates, or 96 in 2-mL vial holders. Add a sample organizer to expand sample capacity for high-throughput and open access environments.

Gradient SmartStart

Easily manage system volume when transferring methods between different LC systems. Automate the timing of pre-injection steps for reduced inject-to-inject cycle times.

TRUE bio-UPLC PERFORMANCE FOR METHOD DEVELOPMENTLeverage the benefits of UPLC Technology to improve your HPLC methods. By scaling your bioseparations methods to UPLC, your assays will benefit from the combination of increased speed, resolution, and sensitivity that comes with modern small particle chemistries.

True UPLC performance comes by pairing a low-dispersion UPLC system with columns containing sub-2-µm particles, with optimized selectivity for the various modes of biochromatography. With our toolbox of simple and intuitive bioseparations applications to help you systematically scale your methods, the process of converting your methods is easy.

Waters offers a series of ACQUITY UPLC columns and chemistries specific for bioseparations. Each of these separations technologies is developed, proven, and quality-control tested for the intended application.

Figure 1. UPLC and ACQUITY UPLC BEH Peptide Separation Technology Columns deliver unparalleled resolution and reproducibility in peptide mapping, here of infliximab (five overlayed chromatograms).

AU

0.00

0.02

0.04

0.06

0.08

0.10

0.12

Retention time (min)10 12 14 16 18 20 22 24 26 28 30 32 34 36 38 40 42

AU

0.00

0.10

0.20

AU

0.02

0.04

0.06

0.08

AU

0.02

0.04

0.06

0.08

0.10

Retention time (min)15 20 25 30 35 40

A

B

C

HPLC

UPLC w/o Auto• Blend

UPLC with Auto• Blend

XBridge® BEH C18 130Å 3.5 µm, 4.6 x 100 mm Mobile phase A: Water Mobile phase B: Acetonitrile Mobile phase C: 1% (v/v) TFA in water

Figure 2. Incorporation of Auto•Blend Technology for peptide mapping produces chromatograms comparable to conventional solvent. Trypsinized infliximab peptide maps were produced using UPLC and HPLC methods. TFA concentration variability was eliminated by providing a stock solution of TFA as its own solvent line.

RELIABLE OPERATION UNDER DEMANDING CONDITIONS Bioseparations assays utilizing IEX, SEC, and HIC modes of biochromatography typically utilize high ionic strength mobile phases containing denaturing agents. The ACQUITY UPLC H-Class Bio System was developed to make biochromatography under such harsh conditions easier and more robust.

The ACQUITY UPLC H-Class Bio system’s bio-inert flow path is made of non-stainless steel and iron-free materials to minimize undesirable protein interactions and maximize system robustness. The result: a system that is ideally suited for routine, trouble-free operation with maximum recovery of biomolecules and consistently high system performance.

BIOCOMPATIBLE MULTI-SOLVENT BLENDING The bioQuaternary Solvent Manager (bioQSM) automatically blends mobile phase combinations and gradients on demand, enabling you to obtain the optimum retention and selectivity requirements for your bioseparation. Designed with bio-inert materials, the bioQSM accommodates the aqueous, high ionic strength mobile phases and organic solvents used in biopharmaceutical applications.

Auto•Blend Plus Technology automatically blends your mobile phase to a specified pH and ionic strength or organic modifier percentage, for an infinite set of analytical conditions. By blending on-line to the desired pH and ionic strength conditions, you can significantly reduce the number of buffers you need to prepare for method development and for routine everyday use.

ACQUITY UPLC BEH450 SEC (4.6 x 300 mm, 2.5 µm)Flow rate = 0.35 mL/minuteInjection volume = 5 µL

SE-HPLC, 450 Å (7.8 x 300 mm, 8 µm)Flow rate = 1.0 mL/minuteInjection volume = 14 µL

Monomer

Dimer

Rs=2.49

Monomer

Dimer

Rs=1.42Figure 3. By combining 450Å sub-3-µm packing materials with a low dispersion ACQUITY UPLC H-Class Bio System, separations with improved resolution and high-throughput of SE-UPLC can be realized for highly aggregated proteins with molecular weights of up to approximately 2000 KDa.

www.waters.com/hclassbio

ACCELERATE YOUR PRODUCTIVITYAdopting a proven analytical technology that improves laboratory efficiency is advantageous for supporting your new projects and products. Designed to support your HPLC, UHPLC, and UPLC methods, the ACQUITY UPLC H-Class Bio System is the ideal solution to address legacy methods, while enabling the transition to true UPLC performance when you are ready.

■■ Reproduce established HPLC methods while enabling seamless transfer to UPLC

■■ Faster methods development with expanded solvent and column capacity and fully scalable column chemistries

Figure 4. Comparison of UPLC and HPLC HILIC separations of RapiFluor-MS® labeled N-glycans from the Waters Glycan Performance Test Standard diluted in water to a concentration of 20 pmole/µL. Injection volumes of 1.2 and 3.7 µL for UPLC and HPLC analyses.

30.00 10.00 12.00 14.00 16.00 18.00 20.00 22.00 24.00 26.00 28.00

64.00 24.00 28.00 32.00 36.00 40.00 44.00 48.00 52.00 56.00 60.00

Alliance® HPLC XBridge Glycan BEH Amide XP, 130Å, 2.5 µm 3.0 mm x 225 mm total length (150 mm + 75 mm)

ACQUITY UPLC H-Class Bio ACQUITY UPLC Glycan BEH Amide, 130Å, 1.7 µm 2.1 mm x 150 mm

Total Run Time = 55 min

Total Run Time = 121.3 min

Fluo

resc

ence

(rel

ativ

e)

Time (min)

1

2

3

4 5

6

7

8

9 10

11

12 13

14 15

16

17 18

19 20 21

Waters, The Science of What’s Possible, Alliance, ACQUITY, Rapi Fluor-MS, UPLC, and XBridge are registered trademarks of Waters Corporation. Auto•Blend Plus is a trademark of Waters Corporation. All other trademarks are the property of their respective owners.

©2017 Waters Corporation. Printed in the U.S.A. August 2017 720005992EN LM-SIG

For your local sales office, please visit www.waters.com/contact

Waters Corporation 34 Maple Street Milford, MA 01757 U.S.A. T: 1 508 478 2000 F: 1 508 872 1990 www.waters.com

www.waters.com/hclassbio

Download the latest version of Bioseparations: Tools, Techniques, and Insights into Biopharmaceutical Analysis