Agilent AdvanceBio Peptide Plus BioLC Columns
Advance Your Confidence for Protein/Peptide Analysis
Reliably Characterize Primary Sequence and Detect Post-Translational Modifications (PTMs)
LC/MS peptide mapping offers simultaneous assessment of critical attributes with high sensitivity and reproducibility. This technique requires a reversed-phase column delivering robust, reproducible results with MS-friendly additives like formic acid.
Confidently identify multiple attributes by LC/MS with new Agilent AdvanceBio Peptide Plus columns, part of the InfinityLab family. Based on Agilent's superficially porous Poroshell technology, these columns feature a hybrid endcapped C-18 stationary phase on a 120 Å pore size, 2.7 µm particle, modified with a charged surface.
These innovative columns enable quick resolution and identification of amino acid modifications in primary structure. They are suitable for both early discovery and transfer to QA/QC, reducing the need for costly, time-consuming method redevelopment.
Key Advantages:
- Quality Assurance: Every column is batch tested with the Agilent peptide standard mix.
- High Sensitivity: Detect critical modifications like oxidation and deamidation even with increasing mass load.
- Formic Acid Compatibility: Multiple geometries allow selection of a single column for early development through QA/QC across various system platforms.
- Charged Surface: Provides sharp, symmetrical peaks using only formic acid as the mobile phase additive.
AdvanceBio Peptide Plus columns are the latest addition to the Agilent AdvanceBio column portfolio, backed by a 60-day warranty. All Agilent BioLC columns are designed for optimum performance with InfinityLab bio-inert LC instruments and supplies.
Quality Assurance: Ensure Confidence in Your Results
Every batch of AdvanceBio Peptide Plus media is tested using the Agilent peptide standard. This mixture includes seven hydrophilic, hydrophobic, and basic peptides, with molecular weights ranging from 757 to 2845 Da. Each column is also tested with a small-molecule probe to ensure efficiency.
The AdvanceBio Peptide Plus columns are an integral part of your characterization workflow, encompassing Sample Prep, Separation, Identification, and Data Analysis.
Workflow Components:
- Sample Prep: The Agilent AssayMAP Bravo protein and peptide sample prep platform offers automation and an optimized workflow for superior efficiency and reproducibility.
- Separation: Enhance accuracy and speed in biomolecule characterization with Agilent AdvanceBio Peptide Plus columns.
- Identification: The Agilent 6545XT AdvanceBio LC/Q-TOF system is designed for multiple biomolecule characterization workflows. Agilent MassHunter software efficiently extracts sample information.
- Data Analysis: Agilent OpenLAB Chromatography Data System and MatchCompare software simplify data review.
Learn more at: www.agilent.com/chem/advancebio
The Solution for Biomolecule Characterization
Combined with Agilent analytical LC/MS instruments, AdvanceBio Peptide Plus columns deliver the speed, sensitivity, and efficiency needed to confirm and identify target proteins and peptides.
LC/MS Conditions for Peptide Identification
| Parameter | Agilent 1290 Infinity II LC | Agilent 6545XT AdvanceBio LC/Q-TOF |
|---|---|---|
| Column | AdvanceBio Peptide Plus 120 Å, 2.1 x 150 mm, 2.7 µm | |
| Mobile phase | A: 0.1% FA, B: 0.1% FA in ACN | |
| Gradient (min) %B | 0 3, 1 3, 31 40, 33 95, 34 95, 34.1 3 | Ion mode: Positive ion mode, dual AJS ESI (profile) |
| Post time | 10 min | Drying gas temperature: 325 °C, Drying gas flow: 13 L/min |
| Injection volume | 1 µL | Sheath gas temperature: 275 °C, Sheath gas flow: 12 L/min |
| Sample thermostat | 5 °C | Nebulizer: 35 psi, Capillary voltage: 4000 V |
| Post time | 5 min | Fragmentor voltage: 175 V, Skimmer voltage: 65 V |
| Column temperature | 55 °C | Oct RF Vpp: 750 V |
| Flow rate | 0.5 mL/min | Acquisition parameters: 2 GHz Extended Dynamic Range, MS mass range 100-1700 m/z, MS/MS mass range 50-1700, MS scan rate: 8 spectra/sec, MS/MS scan rate: 3 spectra/sec |
| Data analysis | Ramped collision energy: 2, Charge state: 3 and >3, Slope: 3.1, Offset: 3.6, BioConfirm software B.08 |
Achieve Accurate Method Reproducibility with LC/MS Peptide Mapping
The precision of peptide mapping critical attributes is demonstrated in replicate injections of a tryptic mAb digest. Arrows indicate peaks selected for %RSD calculations.
| Peak no. | RT | FWHM | Area | Height | %RSD |
|---|---|---|---|---|---|
| 1 | 0.27 | 1.41 | 0.40 | 1.92 | |
| 2 | 0.09 | 0.43 | 0.67 | 0.86 | |
| 3 | 0.06 | 0.73 | 0.59 | 1.10 | |
| 4 | 0.07 | 0.90 | 3.58 | 0.69 | |
| 5 | 0.04 | 0.48 | 0.36 | 0.73 | |
| 6 | 0.05 | 0.82 | 0.67 | 0.41 | |
| 7 | 0.05 | 0.50 | 0.94 | 0.34 | |
| 8 | 0.02 | 0.70 | 0.89 | 1.27 | |
| 9 | 0.03 | 0.40 | 1.18 | 0.39 |
Successfully Separate and Identify a Wide Range of Peptides
The AdvanceBio peptide identification chromatogram displays numerous identified peptides, aiding in comprehensive analysis. (Some identified peptides are not labeled).
Detect Critical PTMs that Occur During Manufacturing, Storage, or Handling
Innovator/Biosimilar Comparison
A comparison of LC/MS peptide maps highlights differences between an innovator and a biosimilar. The highlighted region at 3.73 minutes shows a difference in the SLSLSPGK peptide, potentially due to additional charge (basic) variants in the biosimilar.
Oxidation
The chromatogram demonstrates the separation of nonoxidized and oxidized DLTMISR peptide using an AdvanceBio Peptide Plus column, showing a relative % oxidation of 1.6%.
Deamidation
This overlaid extracted compound chromatogram illustrates the separation of nondeamidated and deamidated peptide using AdvanceBio Peptide Plus columns. Modified sites are marked in red, indicating a total relative % deamidation of 25.52%.
Accurate, Efficient Biopharmaceutical Analysis
Agilent AdvanceBio columns, including the AdvanceBio Peptide Plus, offer consistent performance for protein and mAb characterization. They are designed to enhance accuracy and speed for characterization, SEC aggregation, IEX charge variants, intact mass, primary structure, and impurities profiling.
These columns are suitable for techniques such as reversed-phase, cleaved glycans by hydrophilic interaction, and mAb titer determination by affinity chromatography.
Agilent BioLC Columns Portfolio Overview:
| Application Area | Reversed-Phase | Ion Exchange | Size Exclusion | HILIC | Reversed-Phase | Affinity | Reversed-Phase | Reversed-Phase |
|---|---|---|---|---|---|---|---|---|
| Product Category | Protein identification and impurity profiling | Charged variant analysis | Aggregation analysis | Glycan analysis | Amino acid analysis | Titer determination and purification | Oligonucleotides | Desalting Cartridges |
| Column Types | PLRP-S, AdvanceBio RP-mAb | Bio MAb, Bio IEX (SAX, SCX, WAX, WCX), PL-SAX, PL-SCX | AdvanceBio SEC, Bio SEC-3, Bio SEC-5 | ZORBAX RRHD 300-HILIC | AdvanceBio Peptide Mapping, AdvanceBio Peptide Plus | Bio-Monolith Protein A, Bio-Monolith Protein G | AdvanceBio Oligonucleotides | AdvanceBio Desalting-RP |
| ZORBAX RRHD 300Å 1.8 µm | ProSEC 300S | Bio-Monolith (QA, DEAE, SO3) | Poroshell 300 | ZORBAX 300SB |
Pair with AdvanceBio Columns for Effective Protein/Peptide Analysis
Agilent AdvanceBio columns are part of the Agilent InfinityLab family and are designed to work in harmony with the InfinityLab Bio-inert LC Solution and supplies, providing unmatched bioseparation power and efficiency.
Agilent 1260 Infinity II Bio-inert LC
Protect your biomolecules with a biologically inert flowpath. The Agilent InfinityLab Bio-inert LC Solution enhances confidence in bio-analysis through its corrosion-resistant titanium solvent delivery system and metal-free sample flow path, ensuring biomolecule integrity and increasing efficiency.
Agilent 6545XT AdvanceBio LC/Q-TOF
A new standard for sensitivity and productivity, this system delivers trusted results for intact proteins, peptide sequences, and post-translational modifications. Key features include MassHunter BioConfirm software for intact protein variations, SWARM autotune, mass accuracies within 10 ppm for glycosylated intact proteins, powerful iterative MS/MS for digested samples, and high uptime.
Agilent Supplies: InfinityLab Quick Connect Fittings
These reusable fittings are true "finger-tight" fittings, stable to 1200 bar, requiring no tools for a perfect connection.
Ordering Information
| Description | Part Number |
|---|---|
| AdvanceBio Peptide Plus, 4.6 x 150 mm, 2.7 µm | 671950-902 |
| AdvanceBio Peptide Plus, 3.0 x 150 mm, 2.7 µm | 671950-302 |
| AdvanceBio Peptide Plus, 2.1 x 250 mm, 2.7 µm | 677950-902 |
| AdvanceBio Peptide Plus, 2.1 x 150 mm, 2.7 µm | 675950-902 |
| AdvanceBio Peptide Plus, 2.1 x 50 mm, 2.7 µm | 673950-902 |
| AdvanceBio Peptide Plus, 4.6 mm guard (3/pk) | 820750-932 |
| AdvanceBio Peptide Plus, 3.0 mm guard (3/pk) | 823750-947 |
| AdvanceBio Peptide Plus, 2.1 mm guard (3/pk) | 821725-946 |
| HSA Peptide Standard Mix | G2455-85001 |
All AdvanceBio columns are rigorously tested to ensure reproducibility and are backed by a 60-day warranty.
Learn More
For more information, visit:
- General: www.agilent.com/chem/advancebio
- Customer Centers: www.agilent.com/chem/contactus
Contact Information:
- USA and Canada: 1-800-227-9770, agilent_inquiries@agilent.com
- Europe: info_agilent@agilent.com
- Asia Pacific: inquiry_Isca@agilent.com
Accelerate Productivity and Improve Outcomes
Agilent CrossLab application support helps accelerate productivity and improve outcomes by assisting with method development, instrument repair transitions, and new instrument method transfers, ensuring the latest enhancements are deployed confidently.
Learn more at: www.agilent.com/crosslab
For Research Use Only. Not for use in diagnostic procedures. This information is subject to change without notice. © Agilent Technologies, Inc., 2017. Printed in the USA, May 2, 2017. 5991-7873EN
