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Biomolecule Purification Characterization, and Analyses Catalog
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Biomolecule Purification Characterization, and Analyses Catalog
Product catalog summary
Introduction
Advancements in genomics, proteomics, metabolomics, and molecular biology are revolutionizing disease diagnosis and treatment. Waters, a leader in separation science and analytical biochemistry, offers innovative tools for biomolecule analysis, including HPLC, UHPLC, UPLC, and LC-MS technologies.
Advancements in genomics, proteomics, metabolomics, and molecular biology are revolutionizing disease diagnosis and treatment. Waters, a leader in separation science and analytical biochemistry, offers innovative tools for biomolecule analysis, including HPLC, UHPLC, UPLC, and LC-MS technologies.
Product Offerings
Waters provides a wide range of chemistry and consumables for biomolecule analysis, such as peptide, protein, oligonucleotide, and glycan columns, along with sample preparation kits and analytical standards.
Waters provides a wide range of chemistry and consumables for biomolecule analysis, such as peptide, protein, oligonucleotide, and glycan columns, along with sample preparation kits and analytical standards.
Factors in Column Selection
Key considerations for selecting HPLC, UHPLC, or UPLC columns include particle composition, column length, and particle size, which influence separation quality, resolution, and system back pressure.
Key considerations for selecting HPLC, UHPLC, or UPLC columns include particle composition, column length, and particle size, which influence separation quality, resolution, and system back pressure.
Method Development
Effective bioseparation methods require optimizing factors like eluent composition, temperature, and pH to minimize secondary interactions and enhance separation efficiency.
Effective bioseparation methods require optimizing factors like eluent composition, temperature, and pH to minimize secondary interactions and enhance separation efficiency.
Column and System Compatibility
Ensuring compatibility between columns and LC systems is crucial for optimal performance, with factors like system dispersion and proper connections significantly impacting chromatographic results.
Ensuring compatibility between columns and LC systems is crucial for optimal performance, with factors like system dispersion and proper connections significantly impacting chromatographic results.
Reproducibility and Quality Control
Batch-to-batch and column-to-column reproducibility are essential for reliable results. Waters emphasizes quality control testing with relevant biological standards to ensure consistency.
Batch-to-batch and column-to-column reproducibility are essential for reliable results. Waters emphasizes quality control testing with relevant biological standards to ensure consistency.
Temperature Effects on Protein Separation
Higher temperatures can improve recovery for intact monoclonal antibodies but may not affect selectivity. Some proteins achieve better separation at lower temperatures, highlighting the importance of temperature evaluation in method development.
Higher temperatures can improve recovery for intact monoclonal antibodies but may not affect selectivity. Some proteins achieve better separation at lower temperatures, highlighting the importance of temperature evaluation in method development.
Flow Rate in Reversed-Phase Protein Separations
Lower flow rates enhance resolution without compromising sensitivity, though they increase analysis time. Flow rate adjustments are significant for larger molecules.
Lower flow rates enhance resolution without compromising sensitivity, though they increase analysis time. Flow rate adjustments are significant for larger molecules.
Gradient Slope in IEX Protein Separation
Different salt concentration gradients affect retention time and resolution. Higher salt gradients lead to earlier protein elution, and shallow gradients may require high salt washes for tightly bound proteins.
Different salt concentration gradients affect retention time and resolution. Higher salt gradients lead to earlier protein elution, and shallow gradients may require high salt washes for tightly bound proteins.
Gradient Duration in Reversed-Phase Peptide Separation
Longer gradients improve resolution but increase analysis time and peak volume. Sample complexity influences gradient duration selection.
Longer gradients improve resolution but increase analysis time and peak volume. Sample complexity influences gradient duration selection.
Amino Acid Analysis
Amino acids are crucial for protein composition and metabolic pathways. Accurate analysis is challenging due to diverse chemical properties and the need for derivatization for detection. Waters offers solutions like Pico•Tag, AccQ•Tag, and AccQ•Tag Ultra for precise amino acid analysis using reversed-phase chromatography.
Amino acids are crucial for protein composition and metabolic pathways. Accurate analysis is challenging due to diverse chemical properties and the need for derivatization for detection. Waters offers solutions like Pico•Tag, AccQ•Tag, and AccQ•Tag Ultra for precise amino acid analysis using reversed-phase chromatography.
UPLC Amino Acid Analysis Solution
This solution provides accurate and reproducible amino acid analysis, leveraging Waters' expertise in separation science and derivatization chemistries. It includes UPLC systems, AccQ•Tag Ultra chemistries, and comprehensive support for protein characterization, cell culture monitoring, and nutritional analysis.
This solution provides accurate and reproducible amino acid analysis, leveraging Waters' expertise in separation science and derivatization chemistries. It includes UPLC systems, AccQ•Tag Ultra chemistries, and comprehensive support for protein characterization, cell culture monitoring, and nutritional analysis.
Overview
The document provides a detailed technical overview of various amino acid analysis methods and kits, focusing on the AccQ•Tag Ultra and Pico•Tag methods. It highlights the differences in chemistry, procedures, and applications between these methods, as well as their compatibility with mass spectrometry.
The document provides a detailed technical overview of various amino acid analysis methods and kits, focusing on the AccQ•Tag Ultra and Pico•Tag methods. It highlights the differences in chemistry, procedures, and applications between these methods, as well as their compatibility with mass spectrometry.
AccQ•Tag Ultra Method
This method utilizes Waters 1.7 µm hybrid-silica BEH Technology particles for high column efficiency and resolution. It includes eCord™ Intelligent Chip Technology for tracking column history. The method offers sharper and better-resolved peaks compared to traditional HPLC, enabling faster and more precise analyses. The derivatization reaction involves 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC), which reacts with primary and secondary amines. The method is compatible with electrospray mass spectrometry, allowing for easy identification of amino acids by molecular weight.
This method utilizes Waters 1.7 µm hybrid-silica BEH Technology particles for high column efficiency and resolution. It includes eCord™ Intelligent Chip Technology for tracking column history. The method offers sharper and better-resolved peaks compared to traditional HPLC, enabling faster and more precise analyses. The derivatization reaction involves 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC), which reacts with primary and secondary amines. The method is compatible with electrospray mass spectrometry, allowing for easy identification of amino acids by molecular weight.
AccQ•Tag Method
This method uses the same pre-column derivatization step as the AccQ•Tag Ultra Method but with different reagents and columns. It involves the AccQ•Fluor™ Reagent for derivatizing amines to yield stable, fluorescent adducts. The method is packaged with pre-packaged reagents and documentation for up to 250 analyses.
This method uses the same pre-column derivatization step as the AccQ•Tag Ultra Method but with different reagents and columns. It involves the AccQ•Fluor™ Reagent for derivatizing amines to yield stable, fluorescent adducts. The method is packaged with pre-packaged reagents and documentation for up to 250 analyses.
Pico•Tag Method
A widely-used technique for HPLC amino acid analysis, applicable to various samples including protein hydrolysates and physiological fluids. It relies on the Edman Degradation reaction, forming stable phenylthiocarbamyl (PTC) derivatives. The method allows for the removal of by-products through vacuum drying, providing accurate quantitation of amino acids.
A widely-used technique for HPLC amino acid analysis, applicable to various samples including protein hydrolysates and physiological fluids. It relies on the Edman Degradation reaction, forming stable phenylthiocarbamyl (PTC) derivatives. The method allows for the removal of by-products through vacuum drying, providing accurate quantitation of amino acids.
Ordering Information
The document lists various kits and accessories for amino acid analysis, including part numbers and descriptions for easy ordering. It covers kits for both AccQ•Tag and Pico•Tag methods, detailing the contents and intended applications.
The document lists various kits and accessories for amino acid analysis, including part numbers and descriptions for easy ordering. It covers kits for both AccQ•Tag and Pico•Tag methods, detailing the contents and intended applications.
Glycan and Glycoprotein Analysis
The document briefly mentions glycan and glycoprotein analysis, emphasizing the importance of these analyses in biopharmaceuticals. It highlights Waters' offerings in robust and reproducible analytical methods for glycan analysis.
The document briefly mentions glycan and glycoprotein analysis, emphasizing the importance of these analyses in biopharmaceuticals. It highlights Waters' offerings in robust and reproducible analytical methods for glycan analysis.
Overview
This document provides a comprehensive analysis of glycoprotein and glycopeptide characterization using Waters ACQUITY UPLC Glycoprotein BEH Amide columns. It covers various methods such as intact glycoprotein profiling, subunit analysis, and glycopeptide mapping, which are essential for understanding protein glycosylation.
This document provides a comprehensive analysis of glycoprotein and glycopeptide characterization using Waters ACQUITY UPLC Glycoprotein BEH Amide columns. It covers various methods such as intact glycoprotein profiling, subunit analysis, and glycopeptide mapping, which are essential for understanding protein glycosylation.
Intact Glycoprotein Analysis
The ACQUITY UPLC Glycoprotein BEH Amide column is optimized for resolving glycoforms of intact and digested glycoproteins. It offers high resolution and selectivity, especially for large, released N-glycans. The column operates at elevated temperatures and uses specific mobile-phase additives to enhance solubility and separation efficiency.
The ACQUITY UPLC Glycoprotein BEH Amide column is optimized for resolving glycoforms of intact and digested glycoproteins. It offers high resolution and selectivity, especially for large, released N-glycans. The column operates at elevated temperatures and uses specific mobile-phase additives to enhance solubility and separation efficiency.
Subunit Analysis
Reversed-phase chromatography is used for analyzing protein subunits generated from enzymatic digestion. The document highlights the use of IdeS protease for generating Fc and F(ab')2 fragments from monoclonal antibodies, providing orthogonal and complementary results compared to traditional methods.
Reversed-phase chromatography is used for analyzing protein subunits generated from enzymatic digestion. The document highlights the use of IdeS protease for generating Fc and F(ab')2 fragments from monoclonal antibodies, providing orthogonal and complementary results compared to traditional methods.
Glycopeptide Analysis
HILIC-based separation is emphasized for resolving glycopeptide micro-heterogeneity, which is challenging with reversed-phase methods. This approach effectively characterizes glycan heterogeneity and site occupancy, particularly for O-linked glycans.
HILIC-based separation is emphasized for resolving glycopeptide micro-heterogeneity, which is challenging with reversed-phase methods. This approach effectively characterizes glycan heterogeneity and site occupancy, particularly for O-linked glycans.
LC and MS Conditions
The document details specific LC and MS conditions for various analyses, including column types, mobile phases, temperature settings, and gradient profiles. It also provides ordering information for the necessary columns and standards.
The document details specific LC and MS conditions for various analyses, including column types, mobile phases, temperature settings, and gradient profiles. It also provides ordering information for the necessary columns and standards.
Data Processing
Data processing is conducted using MassLynx and UNIFI software, ensuring accurate mass confirmation and glycan identification.
Data processing is conducted using MassLynx and UNIFI software, ensuring accurate mass confirmation and glycan identification.
Conclusion
The document underscores the versatility and efficiency of the ACQUITY UPLC Glycoprotein BEH Amide column in glycoprotein analysis, offering detailed methodologies and conditions for achieving high-resolution separations and accurate characterizations.
The document underscores the versatility and efficiency of the ACQUITY UPLC Glycoprotein BEH Amide column in glycoprotein analysis, offering detailed methodologies and conditions for achieving high-resolution separations and accurate characterizations.
Overview
The document provides detailed information on the use of various Waters products for glycoprotein and glycan analysis, focusing on the ACQUITY UPLC Glycoprotein BEH Amide Columns and GlycoWorks Sample Preparation Kits. It includes specifications, procedures, and ordering information for these products.
The document provides detailed information on the use of various Waters products for glycoprotein and glycan analysis, focusing on the ACQUITY UPLC Glycoprotein BEH Amide Columns and GlycoWorks Sample Preparation Kits. It includes specifications, procedures, and ordering information for these products.
Specifications
The ACQUITY UPLC Glycoprotein BEH Amide Column is used for separating glycoproteins, specifically ribonuclease A and B from bovine pancreas. The column features a 300 Å pore size and 1.7 µm particle size, ensuring high resolution and reproducibility. The document also details the use of fluorescence detection at specific wavelengths and a column temperature of 45°C.
The ACQUITY UPLC Glycoprotein BEH Amide Column is used for separating glycoproteins, specifically ribonuclease A and B from bovine pancreas. The column features a 300 Å pore size and 1.7 µm particle size, ensuring high resolution and reproducibility. The document also details the use of fluorescence detection at specific wavelengths and a column temperature of 45°C.
Procedures
The document outlines the GlycoWorks RapiFluor-MS N-Glycan Kit, which simplifies the preparation of N-glycan samples. The process involves three main steps: deglycosylation, labeling, and purification, which can be completed in under an hour. The kit is designed for high-throughput processing and is compatible with automation.
The document outlines the GlycoWorks RapiFluor-MS N-Glycan Kit, which simplifies the preparation of N-glycan samples. The process involves three main steps: deglycosylation, labeling, and purification, which can be completed in under an hour. The kit is designed for high-throughput processing and is compatible with automation.
Standards and Calibration
The Glycan Performance Test Standards and Dextran Calibration Ladders are used for benchmarking and method development. These standards help ensure the accuracy and reliability of glycan analysis by providing a means to calibrate systems based on glucose units (GU).
The Glycan Performance Test Standards and Dextran Calibration Ladders are used for benchmarking and method development. These standards help ensure the accuracy and reliability of glycan analysis by providing a means to calibrate systems based on glucose units (GU).
Ordering Information
The document provides part numbers and descriptions for various products, including columns, kits, and standards. It also includes information on additional accessories and reagents necessary for glycan analysis.
The document provides part numbers and descriptions for various products, including columns, kits, and standards. It also includes information on additional accessories and reagents necessary for glycan analysis.
Recommendations
Regular use of the Glycoprotein Performance Test Standard is recommended for monitoring column and system performance. The document emphasizes the importance of using the correct standards and calibration ladders to ensure accurate results.
Regular use of the Glycoprotein Performance Test Standard is recommended for monitoring column and system performance. The document emphasizes the importance of using the correct standards and calibration ladders to ensure accurate results.
Monosaccharide and Sialic Acid Analysis from Glycoproteins
Monosaccharide Analyses
The document outlines the analysis of neutral monosaccharides found in N-linked and O-linked glycans, including N-acetylglucosamine (GlcNAc), N-acetylgalactosamine (GalNAc), galactose (Gal), glucose (Glc), mannose (Man), and fucose (Fuc). The process involves acid hydrolysis using trifluoroacetic acid or hydrochloric acid, followed by derivatization with 2-aminobenzoic acid (2-AA) for high-resolution analysis.
Monosaccharide Analyses
The document outlines the analysis of neutral monosaccharides found in N-linked and O-linked glycans, including N-acetylglucosamine (GlcNAc), N-acetylgalactosamine (GalNAc), galactose (Gal), glucose (Glc), mannose (Man), and fucose (Fuc). The process involves acid hydrolysis using trifluoroacetic acid or hydrochloric acid, followed by derivatization with 2-aminobenzoic acid (2-AA) for high-resolution analysis.
Sialic Acid Analyses
The document describes the analysis of sialic acids, particularly N-acetyl-neuraminic acid (Neu5Ac) and N-glycolyl-neuraminic acid (Neu5Gc), which are important for biopharmaceuticals. The method involves mild acid hydrolysis and derivatization with DMB dye, noting the sensitivity of DMB-labeled sialic acids to light and degradation.
The document describes the analysis of sialic acids, particularly N-acetyl-neuraminic acid (Neu5Ac) and N-glycolyl-neuraminic acid (Neu5Gc), which are important for biopharmaceuticals. The method involves mild acid hydrolysis and derivatization with DMB dye, noting the sensitivity of DMB-labeled sialic acids to light and degradation.
LC Conditions and Column Specifications
The document provides detailed LC conditions for monosaccharide and sialic acid analysis, including mobile phases, flow rates, and column specifications. It highlights the use of ACQUITY UPLC and XBridge BEH C18 columns with varying particle sizes for improved resolution and throughput.
The document provides detailed LC conditions for monosaccharide and sialic acid analysis, including mobile phases, flow rates, and column specifications. It highlights the use of ACQUITY UPLC and XBridge BEH C18 columns with varying particle sizes for improved resolution and throughput.
Effect of Particle Size
The document discusses the impact of particle size on the analysis of 2-AA labeled monosaccharides, noting that smaller particles (1.7 µm) offer higher throughput and improved resolution.
The document discusses the impact of particle size on the analysis of 2-AA labeled monosaccharides, noting that smaller particles (1.7 µm) offer higher throughput and improved resolution.
Ordering Information
Detailed ordering information for various columns is provided, including part numbers and dimensions.
Detailed ordering information for various columns is provided, including part numbers and dimensions.
Oligonucleotide Analysis
The document covers the analysis of oligonucleotides using ion-pair, reversed-phase chromatography. It emphasizes the use of BEH Technology particles for exceptional resolution and column life, and provides LC conditions for the analysis of detritylated oligonucleotides.
The document covers the analysis of oligonucleotides using ion-pair, reversed-phase chromatography. It emphasizes the use of BEH Technology particles for exceptional resolution and column life, and provides LC conditions for the analysis of detritylated oligonucleotides.
UPLC-MS Analysis of Interfering RNA Oligonucleotides
A method for analyzing small interfering RNAs (siRNA) using UPLC-MS is described, highlighting the ability to resolve and characterize oligonucleotide sequences.
A method for analyzing small interfering RNAs (siRNA) using UPLC-MS is described, highlighting the ability to resolve and characterize oligonucleotide sequences.
Conclusion
The document provides comprehensive methodologies for the analysis of monosaccharides, sialic acids, and oligonucleotides, emphasizing the importance of column selection and LC conditions for achieving high-resolution and reliable results.
The document provides comprehensive methodologies for the analysis of monosaccharides, sialic acids, and oligonucleotides, emphasizing the importance of column selection and LC conditions for achieving high-resolution and reliable results.
Overview
The document provides detailed information on the purification and analysis of RNA and DNA using various chromatography techniques, specifically focusing on the use of XBridge Oligonucleotide Columns and Gen-Pak FAX Anion-Exchange Columns. It also discusses the use of Waters ACQUITY UPLC systems for peptide analysis.
The document provides detailed information on the purification and analysis of RNA and DNA using various chromatography techniques, specifically focusing on the use of XBridge Oligonucleotide Columns and Gen-Pak FAX Anion-Exchange Columns. It also discusses the use of Waters ACQUITY UPLC systems for peptide analysis.
Purification and Analysis of RNA
The document describes the successful purification of crude single-stranded RNA using an Alliance System, achieving a purity of approximately 95% with a yield of 55%. XBridge Oligonucleotide Columns are highlighted for their effectiveness in analyzing and purifying siRNA, with specific LC conditions provided for optimal separation.
The document describes the successful purification of crude single-stranded RNA using an Alliance System, achieving a purity of approximately 95% with a yield of 55%. XBridge Oligonucleotide Columns are highlighted for their effectiveness in analyzing and purifying siRNA, with specific LC conditions provided for optimal separation.
Separation of Oligodeoxythymidine Ladder
The document details the separation of 5–25 mer detritylated oligodeoxythymidine using specific LC conditions, including the use of an Alliance HPLC system and XBridge Oligonucleotide BEH C18 columns.
The document details the separation of 5–25 mer detritylated oligodeoxythymidine using specific LC conditions, including the use of an Alliance HPLC system and XBridge Oligonucleotide BEH C18 columns.
Purification of siRNA Duplex
The purification process for siRNA duplexes from impurities is outlined, with specific LC conditions and column specifications provided.
The purification process for siRNA duplexes from impurities is outlined, with specific LC conditions and column specifications provided.
Chromatography of PCR Amplification Mixture
The document describes the separation of a PCR amplification mixture using Gen-Pak FAX columns, detailing the LC conditions and the separation of specific DNA fragments.
The document describes the separation of a PCR amplification mixture using Gen-Pak FAX columns, detailing the LC conditions and the separation of specific DNA fragments.
Separation of DNA Restriction Fragments
The document provides information on the separation of DNA restriction fragments using Gen-Pak FAX columns, with detailed LC conditions and a list of separated compounds.
The document provides information on the separation of DNA restriction fragments using Gen-Pak FAX columns, with detailed LC conditions and a list of separated compounds.
MassPREP Oligonucleotide Standard
The document introduces the MassPREP Oligonucleotide Standard for benchmarking and method development, detailing its composition and use in verifying HPLC/UPLC performance.
The document introduces the MassPREP Oligonucleotide Standard for benchmarking and method development, detailing its composition and use in verifying HPLC/UPLC performance.
Oligonucleotide Desalting
The use of Oasis µElution Plates for oligonucleotide desalting prior to MS analysis is discussed, highlighting their high throughput and low elution volumes.
The use of Oasis µElution Plates for oligonucleotide desalting prior to MS analysis is discussed, highlighting their high throughput and low elution volumes.
Peptide Analysis
The document discusses the challenges of peptide analysis and the use of reversed-phase chromatography for peptide separations. It highlights various column chemistries, including Peptide BEH C18, Peptide CSH C18, and Peptide HSS T3 columns, for different peptide separation needs.
The document discusses the challenges of peptide analysis and the use of reversed-phase chromatography for peptide separations. It highlights various column chemistries, including Peptide BEH C18, Peptide CSH C18, and Peptide HSS T3 columns, for different peptide separation needs.
Overview
The document provides detailed information on various chromatography columns and kits designed for peptide and polypeptide separations. It includes specifications, part numbers, and descriptions of different products offered by Waters, focusing on their applications in analytical and preparative chromatography.
The document provides detailed information on various chromatography columns and kits designed for peptide and polypeptide separations. It includes specifications, part numbers, and descriptions of different products offered by Waters, focusing on their applications in analytical and preparative chromatography.
Specifications
The document lists multiple chromatography columns with varying dimensions and particle sizes, such as BEH C18, CSH C18, and HSS T3, each with specific part numbers. These columns are designed for different applications, including UPLC and HPLC, and are available in various configurations to suit different analytical needs.
The document lists multiple chromatography columns with varying dimensions and particle sizes, such as BEH C18, CSH C18, and HSS T3, each with specific part numbers. These columns are designed for different applications, including UPLC and HPLC, and are available in various configurations to suit different analytical needs.
Product Descriptions
The document describes several product lines, including:
The document describes several product lines, including:
- XBridge Peptide BEH C18: Available in different particle sizes and dimensions, suitable for method validation kits.
- ACQUITY UPLC Peptide CSH C18: Offers columns and kits with specific particle sizes, designed for high-resolution separations.
- XSelect Peptide CSH C18: Includes guards, columns, and kits with various particle sizes for enhanced peptide separations.
- ACQUITY UPLC Peptide HSS T3: Provides columns and kits optimized for peptide analysis with specific particle sizes.
Method Validation Kits
Each kit typically contains multiple columns from different batches to ensure consistency and includes a vial of Cytochrome c Digestion Standard for quality control testing.
Each kit typically contains multiple columns from different batches to ensure consistency and includes a vial of Cytochrome c Digestion Standard for quality control testing.
Applications and Recommendations
The document highlights the use of cation-exchange chromatography for peptide separations, particularly when alternative selectivity is required. It also emphasizes the importance of batch-to-batch reproducibility and the use of specific QC tests to ensure consistent results.
The document highlights the use of cation-exchange chromatography for peptide separations, particularly when alternative selectivity is required. It also emphasizes the importance of batch-to-batch reproducibility and the use of specific QC tests to ensure consistent results.
Additional Products
The document lists various accessories and additional products, such as cartridge holders, replacement O-rings, and method development kits, which support the chromatography process.
The document lists various accessories and additional products, such as cartridge holders, replacement O-rings, and method development kits, which support the chromatography process.
Conclusion
Waters offers a comprehensive range of chromatography products designed to meet the needs of peptide and polypeptide separations, with a focus on high performance, reproducibility, and method validation.
Waters offers a comprehensive range of chromatography products designed to meet the needs of peptide and polypeptide separations, with a focus on high performance, reproducibility, and method validation.
Overview
This document provides detailed information on various HPLC and UHPLC columns and standards used for peptide analysis. It includes specifications, ordering information, and recommended conditions for use.
This document provides detailed information on various HPLC and UHPLC columns and standards used for peptide analysis. It includes specifications, ordering information, and recommended conditions for use.
1. BioSuite C18 Columns
These columns are available in different dimensions and are used for peptide analysis. They are QC tested with tryptic digest of cytochrome c to ensure performance consistency. The document lists part numbers for different sizes and provides eluent conditions, flow rate, gradient, injection volume, temperature, and UV detection settings.
These columns are available in different dimensions and are used for peptide analysis. They are QC tested with tryptic digest of cytochrome c to ensure performance consistency. The document lists part numbers for different sizes and provides eluent conditions, flow rate, gradient, injection volume, temperature, and UV detection settings.
2. MassPREP Peptide Standards
These standards are used for benchmarking, method development, and troubleshooting. They contain a void volume marker and nine peptides with varying polarities and isoelectric points. The document provides part numbers and descriptions for ordering.
These standards are used for benchmarking, method development, and troubleshooting. They contain a void volume marker and nine peptides with varying polarities and isoelectric points. The document provides part numbers and descriptions for ordering.
3. Delta-Pak Columns
These columns are suitable for separating peptides, proteins, and natural products. They are available in different pore sizes and bonded phases. The document includes specifications, ordering information, and conditions for synthetic peptide separation.
These columns are suitable for separating peptides, proteins, and natural products. They are available in different pore sizes and bonded phases. The document includes specifications, ordering information, and conditions for synthetic peptide separation.
4. Symmetry Columns
These columns are designed for peptide separations with high purity and low silanol activity. They offer consistent results and are available in different pore sizes and chemistries. The document provides ordering information and discusses the effects of pore size on peptide selectivity.
These columns are designed for peptide separations with high purity and low silanol activity. They offer consistent results and are available in different pore sizes and chemistries. The document provides ordering information and discusses the effects of pore size on peptide selectivity.
5. BioSuite Cation-Exchange Columns
These columns use sulfopropyl ligand chemistry for peptide isolation based on charge differences. They are available in non-porous and porous forms for different separation needs. The document includes ordering information and conditions for angiotensin separation.
These columns use sulfopropyl ligand chemistry for peptide isolation based on charge differences. They are available in non-porous and porous forms for different separation needs. The document includes ordering information and conditions for angiotensin separation.
6. Additional Peptide Consumables
The document lists various digestion standards and quantitative peptide standards for use in proteomics and LC-MS systems. It provides descriptions and part numbers for ordering.
The document lists various digestion standards and quantitative peptide standards for use in proteomics and LC-MS systems. It provides descriptions and part numbers for ordering.
SILAC Hi3 Phos B Standard 186007083
The SILAC Hi3 Phos B standard is derived from the top six ionizing peptides of rabbit phosphorylase B protein, similar to its non-labeled counterpart. The key distinction is the inclusion of a heavy labeled reference on the lysine (K) or arginine (R) end of the peptide.
The SILAC Hi3 Phos B standard is derived from the top six ionizing peptides of rabbit phosphorylase B protein, similar to its non-labeled counterpart. The key distinction is the inclusion of a heavy labeled reference on the lysine (K) or arginine (R) end of the peptide.
SILAC Hi3 E. coli Standard 186007084
This standard is formulated from the top six ionizing peptides of the E. coli ClpB protein, with a heavy labeled reference on the lysine (K) or arginine (R) end, differentiating it from its non-labeled version.
This standard is formulated from the top six ionizing peptides of the E. coli ClpB protein, with a heavy labeled reference on the lysine (K) or arginine (R) end, differentiating it from its non-labeled version.
Quantitative Peptide Retention Standard 186006555
This standard aids in chromatographic separations, ensuring result confidence. It features peak retention for reproducibility, UV absorptivity, a wide mass range for MS, water solubility, and tryptic-like peptides for mapping studies.
This standard aids in chromatographic separations, ensuring result confidence. It features peak retention for reproducibility, UV absorptivity, a wide mass range for MS, water solubility, and tryptic-like peptides for mapping studies.
MassPREP Phosphopeptide Standards
These standards offer control over sample preparation, allowing the use of pure peptides or defining phosphopeptide to unmodified peptide ratios. They include various kits and mixtures for optimizing phosphopeptide detection in LC-MS, LC/UV, and MALDI-MS.
These standards offer control over sample preparation, allowing the use of pure peptides or defining phosphopeptide to unmodified peptide ratios. They include various kits and mixtures for optimizing phosphopeptide detection in LC-MS, LC/UV, and MALDI-MS.
RapiGest SF Protein Digestion Surfactant
RapiGest SF enhances protein enzymatic digestions by solubilizing and unfolding proteins, making them more accessible to proteases without denaturing them. It is available in various vial sizes.
RapiGest SF enhances protein enzymatic digestions by solubilizing and unfolding proteins, making them more accessible to proteases without denaturing them. It is available in various vial sizes.
Protein Analysis
Waters provides solutions for protein separation and characterization using techniques like reversed-phase, hydrophilic-interaction, SEC, and ion-exchange. These methods are crucial for developing biopharmaceuticals and diagnostic reagents.
Waters provides solutions for protein separation and characterization using techniques like reversed-phase, hydrophilic-interaction, SEC, and ion-exchange. These methods are crucial for developing biopharmaceuticals and diagnostic reagents.
Intact Protein and mAb Subunit Analysis
The BioResolve RP mAb Polyphenyl Columns are designed for high-quality LC or LC-MS analyses of intact monoclonal antibodies and subunits. They offer improved resolution, quantitation accuracy, and enhanced MS data, with compatibility across different chromatography systems.
The BioResolve RP mAb Polyphenyl Columns are designed for high-quality LC or LC-MS analyses of intact monoclonal antibodies and subunits. They offer improved resolution, quantitation accuracy, and enhanced MS data, with compatibility across different chromatography systems.
Introduction
This document provides a detailed overview of various chromatography columns and standards used for the characterization of proteins, monoclonal antibodies (mAbs), and antibody-drug conjugates (ADCs). It highlights the use of high-resolution chromatography-mass spectrometry and the benefits of specific column technologies.
This document provides a detailed overview of various chromatography columns and standards used for the characterization of proteins, monoclonal antibodies (mAbs), and antibody-drug conjugates (ADCs). It highlights the use of high-resolution chromatography-mass spectrometry and the benefits of specific column technologies.
Specifications and Features
The BioResolve RP mAb Polyphenyl Column is noted for its solid-core particle design and innovative polyphenyl ligand bonding, which minimizes method transfer concerns. It is suitable for use throughout discovery, product development, and QC manufacturing controls. The document also details the Waters mAb Subunit Standard, which is used for benchmarking and method development.
The BioResolve RP mAb Polyphenyl Column is noted for its solid-core particle design and innovative polyphenyl ligand bonding, which minimizes method transfer concerns. It is suitable for use throughout discovery, product development, and QC manufacturing controls. The document also details the Waters mAb Subunit Standard, which is used for benchmarking and method development.
Analytical Techniques
The document discusses the use of reversed-phase HPLC and UPLC for protein separation, emphasizing the high-resolution analysis capabilities of Waters' BEH Technology Protein Separation Columns. These columns are designed to handle proteins of various sizes, hydrophobicities, and isoelectric points, and are tested for batch-to-batch reproducibility.
The document discusses the use of reversed-phase HPLC and UPLC for protein separation, emphasizing the high-resolution analysis capabilities of Waters' BEH Technology Protein Separation Columns. These columns are designed to handle proteins of various sizes, hydrophobicities, and isoelectric points, and are tested for batch-to-batch reproducibility.
Applications
The BioResolve column is praised for its ability to distinguish between closely related species, such as wild-type nanobodies and their modified forms. This capability is crucial for analyzing antibody modifications.
The BioResolve column is praised for its ability to distinguish between closely related species, such as wild-type nanobodies and their modified forms. This capability is crucial for analyzing antibody modifications.
Ordering Information
Detailed ordering information is provided for various column dimensions and method validation kits, including part numbers and descriptions for BioResolve RP mAb Polyphenyl Columns and Protein BEH C4 Columns.
Detailed ordering information is provided for various column dimensions and method validation kits, including part numbers and descriptions for BioResolve RP mAb Polyphenyl Columns and Protein BEH C4 Columns.
Protein Standards
The MassPREP Protein Standard Mix is highlighted as a benchmarking tool for validating column and LC system performance. It includes proteins with a range of molecular weights and isoelectric points.
The MassPREP Protein Standard Mix is highlighted as a benchmarking tool for validating column and LC system performance. It includes proteins with a range of molecular weights and isoelectric points.
Hydrophobic Interaction Chromatography (HIC)
The document introduces Protein-Pak Hi Res HIC Columns, which are designed for non-denaturing protein characterization. These columns use non-porous particles for efficient separations and are suitable for high-throughput needs.
The document introduces Protein-Pak Hi Res HIC Columns, which are designed for non-denaturing protein characterization. These columns use non-porous particles for efficient separations and are suitable for high-throughput needs.
Conclusion
This document serves as a comprehensive guide for selecting and using chromatography columns and standards for protein analysis, offering insights into method development, troubleshooting, and quality control.
This document serves as a comprehensive guide for selecting and using chromatography columns and standards for protein analysis, offering insights into method development, troubleshooting, and quality control.
Overview
This document provides detailed information on the use of various chromatography columns and techniques for the analysis and separation of proteins, peptides, and antibody drug conjugates (ADCs). It includes specifications, procedures, and recommendations for different types of columns and their applications.
This document provides detailed information on the use of various chromatography columns and techniques for the analysis and separation of proteins, peptides, and antibody drug conjugates (ADCs). It includes specifications, procedures, and recommendations for different types of columns and their applications.
1. Protein-Pak Hi Res HIC Column
- Used for the separation of cysteine-based ADCs using hydrophobic interaction chromatography (HIC).
- Conditions include a gradient of decreasing salt concentration with non-denaturing eluents.
- Specifications: 2.5 µm, 4.6 × 100 mm column, flow rate of 0.7 mL/min, and UV detection at 280 nm.
2. BioSuite Phenyl HIC Columns
- Designed for hydrophobic-interaction chromatography, providing an alternative to reversed-phase chromatography (RPC).
- Characterized by adsorption to a weakly hydrophobic surface at high salt concentrations.
- Specifications: 1000 Å, 10 µm column, flow rate of 1.0 mL/min, and UV detection at 280 nm.
3. ACQUITY UPLC Glycoprotein BEH Amide Columns
- Used for profiling IgG subunit glycoforms and middle-up/middle-down analysis of mAbs.
- Combines IdeS digestion with reversed-phase chromatography for mAb characterization.
- Specifications: 300 Å, 1.7 µm column, flow rate of 0.2 mL/min, and UV detection at 220 nm.
4. Lifetime Testing
- Demonstrates the robustness of the ACQUITY UPLC Glycoprotein BEH Amide column over 300 sequential injections.
- Monitors parameters such as retention time, resolution, and system pressure.
5. Ordering Information
- Includes part numbers and dimensions for various columns and standards.
- Emphasizes the importance of quality control testing for consistency in chromatography results.
Aggregate Analysis
The ACQUITY UPLC Technology significantly enhances laboratory productivity by enabling faster and more efficient analysis compared to traditional HPLC methods. The ACQUITY UPLC SEC System Solution, utilizing ethylene-bridged-hybrid (BEH) diol-coated particle technology, allows for rapid determination of aggregation levels in therapeutic monoclonal antibodies, reducing analysis time by up to 10 times. The system's optimized column chemistry minimizes the need for high salt concentration mobile phases, ensuring consistent and reliable results.
The ACQUITY UPLC Technology significantly enhances laboratory productivity by enabling faster and more efficient analysis compared to traditional HPLC methods. The ACQUITY UPLC SEC System Solution, utilizing ethylene-bridged-hybrid (BEH) diol-coated particle technology, allows for rapid determination of aggregation levels in therapeutic monoclonal antibodies, reducing analysis time by up to 10 times. The system's optimized column chemistry minimizes the need for high salt concentration mobile phases, ensuring consistent and reliable results.
BEH Technology
BEH Technology represents a second-generation hybrid particle structure that combines the best properties of inorganic and organic packings. This technology provides superior mechanical strength, efficiency, and stability, enabling high-speed, sensitive, and high-resolution UPLC separations for both small and large molecules.
BEH Technology represents a second-generation hybrid particle structure that combines the best properties of inorganic and organic packings. This technology provides superior mechanical strength, efficiency, and stability, enabling high-speed, sensitive, and high-resolution UPLC separations for both small and large molecules.
Separation of Protein and Peptide Standards
The ACQUITY UPLC Protein BEH SEC columns are available in various pore sizes (125 Å, 200 Å, and 450 Å) to accommodate different molecular weight ranges. These columns are designed to improve the resolution and speed of protein and peptide separations, offering significant advantages over traditional methods.
The ACQUITY UPLC Protein BEH SEC columns are available in various pore sizes (125 Å, 200 Å, and 450 Å) to accommodate different molecular weight ranges. These columns are designed to improve the resolution and speed of protein and peptide separations, offering significant advantages over traditional methods.
SEC Analysis of Insulin
Size-exclusion chromatography (SEC) is the standard method for analyzing insulin and its aggregates. The ACQUITY UPLC BEH SEC columns provide improved resolution and reduced analysis time compared to traditional HPLC-based SEC methods, enhancing the efficiency of insulin analysis.
Size-exclusion chromatography (SEC) is the standard method for analyzing insulin and its aggregates. The ACQUITY UPLC BEH SEC columns provide improved resolution and reduced analysis time compared to traditional HPLC-based SEC methods, enhancing the efficiency of insulin analysis.
LC-MS Analysis of Monoclonal Antibodies
Using non-denaturing conditions, the ACQUITY UPLC BEH SEC columns allow for detailed analysis of monoclonal antibodies, including aggregate and monomer detection. The technology provides accurate mass determination and helps identify unexpected peaks, facilitating better characterization and quality control.
Using non-denaturing conditions, the ACQUITY UPLC BEH SEC columns allow for detailed analysis of monoclonal antibodies, including aggregate and monomer detection. The technology provides accurate mass determination and helps identify unexpected peaks, facilitating better characterization and quality control.
Reduced Requirement for High Salt Concentration
The unique surface chemistry of the ACQUITY UPLC Protein BEH SEC columns reduces secondary ionic interactions, allowing for lower salt concentrations in mobile phases. This results in improved peak shapes and reduced retention times, particularly for basic proteins like lysozyme.
The unique surface chemistry of the ACQUITY UPLC Protein BEH SEC columns reduces secondary ionic interactions, allowing for lower salt concentrations in mobile phases. This results in improved peak shapes and reduced retention times, particularly for basic proteins like lysozyme.
Column Stability and Performance
BEH particle technology, combined with innovative diol-bonding processes, ensures exceptional column stability and performance. This technology extends column life and enhances the reliability of size-exclusion chromatography, making it suitable for a wide range of biomolecular analyses.
BEH particle technology, combined with innovative diol-bonding processes, ensures exceptional column stability and performance. This technology extends column life and enhances the reliability of size-exclusion chromatography, making it suitable for a wide range of biomolecular analyses.
Specifications
- Flow rate: 1.0 mL/min
- Column temperature: 25 °C
- UV detection: 220 nm
- Available columns: BioSuite 4.6 mm, 7.8 mm, 7.5 mm analytical and 21.5 mm preparative columns.
- Flow rate: 1.0 mL/min
- Column temperature: 25 °C
- UV detection: 220 nm
- Available columns: BioSuite 4.6 mm, 7.8 mm, 7.5 mm analytical and 21.5 mm preparative columns.
Column Specifications
- BioSuite 125 Å: Globular Protein MW Range 5000–150,000, Branched Dextrans 1000–30,000, Linear PEG/PEO 500–15,000
- BioSuite 250 Å: Globular Protein MW Range 10,000–500,000, Branched Dextrans 2000–70,000, Linear PEG/PEO 1000–35,000
- BioSuite 450 Å: Globular Protein MW Range 20,000–1,000,000, Branched Dextrans 4000–500,000, Linear PEG/PEO 2000–250,000
- BioSuite 125 Å: Globular Protein MW Range 5000–150,000, Branched Dextrans 1000–30,000, Linear PEG/PEO 500–15,000
- BioSuite 250 Å: Globular Protein MW Range 10,000–500,000, Branched Dextrans 2000–70,000, Linear PEG/PEO 1000–35,000
- BioSuite 450 Å: Globular Protein MW Range 20,000–1,000,000, Branched Dextrans 4000–500,000, Linear PEG/PEO 2000–250,000
Protein-Pak and Shodex Size-Exclusion HPLC Columns
- Protein-Pak packings are based on a 10 µm, diol-bonded silica.
- Shodex offers 7 µm, high-resolution, gel filtration packings.
- Resolution depends more on sample mass and volume than interaction with the stationary phase.
- Secondary interactions can be reduced by increasing ionic strength or adding methanol.
- Protein-Pak packings are based on a 10 µm, diol-bonded silica.
- Shodex offers 7 µm, high-resolution, gel filtration packings.
- Resolution depends more on sample mass and volume than interaction with the stationary phase.
- Secondary interactions can be reduced by increasing ionic strength or adding methanol.
Standard Protein Mix on KW-803 Column
- Column: Shodex KW-803, 7 µm, 8 × 300 mm
- Eluent: 25 mM sodium phosphate, pH 6.8
- Flow rate: 0.72 mL/min
- UV detection: 280 nm
- Compounds: Blue dextran, Ferritin, Aldolase, Bovine serum albumin
- Column: Shodex KW-803, 7 µm, 8 × 300 mm
- Eluent: 25 mM sodium phosphate, pH 6.8
- Flow rate: 0.72 mL/min
- UV detection: 280 nm
- Compounds: Blue dextran, Ferritin, Aldolase, Bovine serum albumin
BioSuite SEC HPLC and UHPLC Columns
- Various specifications for different pore sizes and particle sizes.
- Suggested volume load for maximum multicomponent resolution varies by column type.
- Various specifications for different pore sizes and particle sizes.
- Suggested volume load for maximum multicomponent resolution varies by column type.
Ion-Exchange Analysis
- Ion-Exchange (IEX) separations use gradients of increasing salt or changing pH.
- BioResolve SCX mAb Columns are used for charge variant profiling of monoclonal antibodies.
- Benefits include high mechanical strength and chemical tolerance.
- Ion-Exchange (IEX) separations use gradients of increasing salt or changing pH.
- BioResolve SCX mAb Columns are used for charge variant profiling of monoclonal antibodies.
- Benefits include high mechanical strength and chemical tolerance.
VanGuard FIT Cartridge
- Designed to protect columns from fouling and extend column life.
- Can be easily replaced to restore column performance.
- Designed to protect columns from fouling and extend column life.
- Can be easily replaced to restore column performance.
BioResolve CX pH Buffers
- Facilitate high-resolution separations with BioResolve SCX mAb Columns.
- Offer a more universal pH gradient applicable to many samples.
- Facilitate high-resolution separations with BioResolve SCX mAb Columns.
- Offer a more universal pH gradient applicable to many samples.
Introduction
This document outlines a pH gradient-based method for charge variant analysis of monoclonal antibodies (mAbs) using BioResolve SCX mAb Columns. The method involves using carefully formulated concentrates to prepare mobile phases with controlled pH and ionic strength, facilitating robust cation-exchange separations.
This document outlines a pH gradient-based method for charge variant analysis of monoclonal antibodies (mAbs) using BioResolve SCX mAb Columns. The method involves using carefully formulated concentrates to prepare mobile phases with controlled pH and ionic strength, facilitating robust cation-exchange separations.
Specifications
The BioResolve SCX mAb Columns are available in various dimensions and are designed for high-resolution ion-exchange chromatography. The columns use a 10x concentrated solution that can be diluted to create a mobile phase with a pH range from 5.0 to 10.2.
The BioResolve SCX mAb Columns are available in various dimensions and are designed for high-resolution ion-exchange chromatography. The columns use a 10x concentrated solution that can be diluted to create a mobile phase with a pH range from 5.0 to 10.2.
Procedures
The document describes the use of VanGuard FIT Cartridges to extend the life of BioResolve SCX mAb Columns by replacing them when particulate fouling occurs. It also details the separation of mAb charge variants using a linear pH gradient on an ACQUITY UPLC H-Class Bio System.
The document describes the use of VanGuard FIT Cartridges to extend the life of BioResolve SCX mAb Columns by replacing them when particulate fouling occurs. It also details the separation of mAb charge variants using a linear pH gradient on an ACQUITY UPLC H-Class Bio System.
Standards and Recommendations
Protein-Pak Hi Res Ion-Exchange Columns are recommended for characterizing recombinant proteins and mAbs. These columns offer high compound binding capacity and are QC tested to ensure consistent performance. The document also provides ordering information for various columns and standards.
Protein-Pak Hi Res Ion-Exchange Columns are recommended for characterizing recombinant proteins and mAbs. These columns offer high compound binding capacity and are QC tested to ensure consistent performance. The document also provides ordering information for various columns and standards.
Data and Analysis
Chromatograms and pH traces are provided to illustrate the separation of mAb charge variants. The document includes data on the reproducibility of Protein-Pak Hi Res IEX Columns across different batches, ensuring consistent performance.
Chromatograms and pH traces are provided to illustrate the separation of mAb charge variants. The document includes data on the reproducibility of Protein-Pak Hi Res IEX Columns across different batches, ensuring consistent performance.
Conclusion
The document emphasizes the importance of using high-quality, consistent columns for the successful development and commercialization of biotherapeutics. It highlights the benefits of using ACQUITY UPLC Technology for enhanced resolution, sensitivity, and throughput in biotherapeutic characterization.
The document emphasizes the importance of using high-quality, consistent columns for the successful development and commercialization of biotherapeutics. It highlights the benefits of using ACQUITY UPLC Technology for enhanced resolution, sensitivity, and throughput in biotherapeutic characterization.
Overview
The document provides detailed information on various HPLC columns and ion-exchange packings used for the separation and purification of complex biocompound mixtures. It includes specifications, ordering information, and guidelines for use.
The document provides detailed information on various HPLC columns and ion-exchange packings used for the separation and purification of complex biocompound mixtures. It includes specifications, ordering information, and guidelines for use.
1. BioSuite IEX HPLC Columns
These columns are designed for ion-exchange chromatography, featuring different matrices, pore sizes, and protein binding capacities. Key specifications include pore size, exclusion limit, and column dimensions. The document emphasizes not exceeding 20% of the column's protein binding capacity for optimal resolution.
These columns are designed for ion-exchange chromatography, featuring different matrices, pore sizes, and protein binding capacities. Key specifications include pore size, exclusion limit, and column dimensions. The document emphasizes not exceeding 20% of the column's protein binding capacity for optimal resolution.
2. Protein-Pak PW HPLC Column Series
This section describes polymeric ion-exchangers available in both glass and steel columns, suitable for HPLC and FPLC systems. The columns are available in various dimensions and are used for both analytical and preparative purposes.
This section describes polymeric ion-exchangers available in both glass and steel columns, suitable for HPLC and FPLC systems. The columns are available in various dimensions and are used for both analytical and preparative purposes.
3. Protein-Pak High Resolution (HR) Ion-Exchange Glass Columns
These columns use polymethacrylate particles with large pores, reducing non-specific adsorption and ensuring high protein recovery. They are compatible with a wide pH range and withstand caustic cleaning solutions. The document details the types of ion exchangers available and their respective properties.
These columns use polymethacrylate particles with large pores, reducing non-specific adsorption and ensuring high protein recovery. They are compatible with a wide pH range and withstand caustic cleaning solutions. The document details the types of ion exchangers available and their respective properties.
4. Advanced Purification (AP) Glass Columns
AP Glass Columns are biocompatible and can be used with various packing materials. They feature adjustable bed heights and are compatible with both analytical and preparative systems. The document lists available sizes and accessories.
AP Glass Columns are biocompatible and can be used with various packing materials. They feature adjustable bed heights and are compatible with both analytical and preparative systems. The document lists available sizes and accessories.
5. AccellPlus Ion-Exchange Packings
These are silica-based materials used for protein sample preparation. They are available as strong anion or weak cation exchangers and are suitable for high flow rates during cleaning cycles. The document provides guidelines for estimating packed bed volume and using Sep-Pak cartridges for sample preparation.
These are silica-based materials used for protein sample preparation. They are available as strong anion or weak cation exchangers and are suitable for high flow rates during cleaning cycles. The document provides guidelines for estimating packed bed volume and using Sep-Pak cartridges for sample preparation.
6. Protein-Pak Affinity Columns
These columns feature epoxy-activated packing for immobilizing ligands via covalent linkage. They are suitable for method screening and small to large-scale separations.
These columns feature epoxy-activated packing for immobilizing ligands via covalent linkage. They are suitable for method screening and small to large-scale separations.
Key Recommendations
For best resolution, do not exceed 20% of the column's protein binding capacity. Use appropriate buffers and flow rates as specified for each column type to ensure optimal performance.
For best resolution, do not exceed 20% of the column's protein binding capacity. Use appropriate buffers and flow rates as specified for each column type to ensure optimal performance.
Packed Bed Volume Estimation
The packed bed volume for Protein-Pak Affinity Epoxy-Activated packing can be estimated by multiplying the weight of the packing used (in grams) by 2 to get the volume in milliliters.
The packed bed volume for Protein-Pak Affinity Epoxy-Activated packing can be estimated by multiplying the weight of the packing used (in grams) by 2 to get the volume in milliliters.
Purification of Carbonic Anhydrase
The purification process involves using a ligand, Sulfanilamide, at 180 µmol/g. The column used is an AP-1 Glass with dimensions 10 × 100 mm. Two buffers are used: Buffer A (100 mM tris sulfate with 200 mM sodium sulfate, pH 8.7) and Buffer B (200 mM potassium thiocyanate in 50 mM tris sulfate, pH 6.5). The gradient profile involves a flow rate of 2 mL/min with UV detection at 280 nm. A protein load of 24 mg of bovine hemolysate yields 0.6 mg of carbonic anhydrase with 100% recovery of esterase activity.
The purification process involves using a ligand, Sulfanilamide, at 180 µmol/g. The column used is an AP-1 Glass with dimensions 10 × 100 mm. Two buffers are used: Buffer A (100 mM tris sulfate with 200 mM sodium sulfate, pH 8.7) and Buffer B (200 mM potassium thiocyanate in 50 mM tris sulfate, pH 6.5). The gradient profile involves a flow rate of 2 mL/min with UV detection at 280 nm. A protein load of 24 mg of bovine hemolysate yields 0.6 mg of carbonic anhydrase with 100% recovery of esterase activity.
Protein-Pak Affinity Columns
These columns are available in various particle sizes and pore sizes, with specific part numbers for ordering. The document provides details on the specifications and ordering information for different types of Protein-Pak Affinity columns.
These columns are available in various particle sizes and pore sizes, with specific part numbers for ordering. The document provides details on the specifications and ordering information for different types of Protein-Pak Affinity columns.
Reversed-Phase Chromatography (RPC)
BioSuite pC18 and pPhenyl RPC columns are used for the separation of proteins, peptides, and other biomolecules. The columns are available in different pore sizes to accommodate proteins of varying sizes. The document details the specifications and applications of these columns, including their suitability for lab-scale isolations and narrow-bore HPLC and LC-MS applications.
BioSuite pC18 and pPhenyl RPC columns are used for the separation of proteins, peptides, and other biomolecules. The columns are available in different pore sizes to accommodate proteins of varying sizes. The document details the specifications and applications of these columns, including their suitability for lab-scale isolations and narrow-bore HPLC and LC-MS applications.
LC-MS Analysis
The document describes the conditions for LC-MS analysis of a reduced monoclonal IgG1 antibody using a BioSuite pPhenyl RPC column. It includes details on the sample, column, eluents, flow rate, gradient, temperature, and detection method.
The document describes the conditions for LC-MS analysis of a reduced monoclonal IgG1 antibody using a BioSuite pPhenyl RPC column. It includes details on the sample, column, eluents, flow rate, gradient, temperature, and detection method.
Nano- and Micro-Flow LC-MS
This section discusses the advantages of nano- and micro-flow LC-MS, including enhanced sensitivity and reduced sample requirements. It highlights the benefits of using smaller column diameters and flow rates to increase sensitivity and reduce matrix effects.
This section discusses the advantages of nano- and micro-flow LC-MS, including enhanced sensitivity and reduced sample requirements. It highlights the benefits of using smaller column diameters and flow rates to increase sensitivity and reduce matrix effects.
iKey Separation Device
The iKey Separation Device is a component of the ionKey/MS System, designed to simplify micro-flow LC-MS analyses. It integrates fluid connections, electronics, and column heating, providing robust and reproducible performance.
The iKey Separation Device is a component of the ionKey/MS System, designed to simplify micro-flow LC-MS analyses. It integrates fluid connections, electronics, and column heating, providing robust and reproducible performance.
Overview
This document provides detailed information on LC-MS analyses designed for low-dispersion nano-UPLC systems, focusing on separation and trapping columns, nanoEase M/Z columns with ZenFit Connection Technology, and various accessories and components used in these systems.
This document provides detailed information on LC-MS analyses designed for low-dispersion nano-UPLC systems, focusing on separation and trapping columns, nanoEase M/Z columns with ZenFit Connection Technology, and various accessories and components used in these systems.
Separation Columns
Separation columns are designed for nano- and microscale separations with MS detection under UPLC conditions at 15,000 psi. They utilize sub-2-µm particle technology for enhanced separation power. Columns range from 75 to 300 µm I.D., supporting flow rates between 200 nL/min and 100 µL/min, allowing flexibility in complex LC-MS analyses.
Separation columns are designed for nano- and microscale separations with MS detection under UPLC conditions at 15,000 psi. They utilize sub-2-µm particle technology for enhanced separation power. Columns range from 75 to 300 µm I.D., supporting flow rates between 200 nL/min and 100 µL/min, allowing flexibility in complex LC-MS analyses.
Trapping Columns
Trapping columns are used to desalt and enrich samples before final separation with MS detection. They are packed with larger 5 µm particles for fast loading and reduced cycle time.
Trapping columns are used to desalt and enrich samples before final separation with MS detection. They are packed with larger 5 µm particles for fast loading and reduced cycle time.
nanoEase M/Z Columns with ZenFit Connection Technology
These columns feature easy-to-use, reusable, fingertight liquid-line connectors, eliminating dead volume and variability. They are compatible with the ACQUITY UPLC M-Class System when equipped with the appropriate upgrade kit.
These columns feature easy-to-use, reusable, fingertight liquid-line connectors, eliminating dead volume and variability. They are compatible with the ACQUITY UPLC M-Class System when equipped with the appropriate upgrade kit.
Ordering Information
The document lists various nanoEase M/Z Peptide, Protein, and HSS Columns, along with their dimensions, particle sizes, and part numbers.
The document lists various nanoEase M/Z Peptide, Protein, and HSS Columns, along with their dimensions, particle sizes, and part numbers.
ACQUITY UPLC M-Class with HDX Technology
This section discusses the use of hydrogen-deuterium exchange mass spectrometry (HDX-MS) for studying protein structural dynamics. The system can quantify small changes in protein conformation and includes the Waters Enzymate™ BEH Pepsin Column for online protein digestion.
This section discusses the use of hydrogen-deuterium exchange mass spectrometry (HDX-MS) for studying protein structural dynamics. The system can quantify small changes in protein conformation and includes the Waters Enzymate™ BEH Pepsin Column for online protein digestion.
LC-MS Accessories
TruView LCMS Certified Vials are highlighted for their low polar analyte adsorption and high recovery of analytes. Waters Certified Containers are also mentioned for their ultra-clean properties and prevention of contamination.
TruView LCMS Certified Vials are highlighted for their low polar analyte adsorption and high recovery of analytes. Waters Certified Containers are also mentioned for their ultra-clean properties and prevention of contamination.
pH Buffers
The document lists pH buffers that are traceable to NIST SRMs, mercury-free, and stable for at least one year, along with their ordering information.
The document lists pH buffers that are traceable to NIST SRMs, mercury-free, and stable for at least one year, along with their ordering information.
Conclusion
The document provides comprehensive ordering information for various components and accessories used in LC-MS analyses, emphasizing the importance of high-quality materials and technologies for accurate and efficient biomarker discovery and protein characterization.
The document provides comprehensive ordering information for various components and accessories used in LC-MS analyses, emphasizing the importance of high-quality materials and technologies for accurate and efficient biomarker discovery and protein characterization.
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Catalog excerpts
omolecule Purification, + Characterization, ar.d^^ Analyses Catalog * INNOVATIVE HPLC, UHPLC, AND UPLC CHEMISTRY CcSSm JLES FOR B!Osi>ARATIONS
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Biomolecule Purification, Characterization, and Analysis Innovative Technologies from the Leader in Separation Science and Analytical Biochemistry Advances in the areas of genomics, proteomics, metabolomics, and molecular and system biology continue to revolutionize the diagnosis and treatment of diseases and increase our fundamental understanding of biological processes. As a leading analytical supplier of instrumentation, software, service and support, and chemistry products, Waters is uniquely positioned to provide researchers the tools, technologies, and integrated solutions desired to tackle...
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Factors to Consider when Investing and Using HPLC, UHPLC, or UPLC Columns for Bioseparations 2 Part 1: Column Selection and Installation 5 Part 2: Bioseparation Method Development Amino Acid Analysis Glycan and Glycoprotein Analysis Oligonucleotide Analysis Peptide Analysis Protein Analysis Nano-Flow and Micro-Flow LC-MS
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Factors to Consider when Investing and Using HPLC, UHPLC, or UPLC Columns for Bioseparations Many factors can affect the quality of data obtained from LC-based separations of peptides, proteins, and other biomolecules. The following pages list just a few of the important factors to consider when selecting an appropriate HPLC, UHPLC, or UPLC column for analytical or lab-scale applications. Once an appropriate column is selected, time must be invested in developing a satisfactory separation, so we have also included a few useful method development “tips and tricks”. We hope that these few examples...
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Effect of Particle Size on SEC Protein Separations ■■Well-packed columns containing small particles can improve a separation ■■System back pressure will increase as particle size decreases ■■Consequently, LC instrumentation can limit potential column use 0.18 A comparison of separations of Waters BEH450 SEC Protein Standard Mix (p/n: 186006842) and Intact mAb Mass Check Standard (p/n: 186006552, diluted to 1 mg/mL) on 450 Å, silica-based 8 µm (Frames A and C) and 450 Å, BEH 3.5 µm (Frames B and D) SEC columns. Both columns were the same dimensions (7.8 x 300 mm) and separations were performed...
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Importance of Making Good Connections from Column to LC System ■■Poor column-to-instrument connections can degrade a chromatographic separation ■■Perceived column leaking can also be caused by a poor connection Sources of Band Spreading – Improper Column Connection Improper Connection Dead/void volume Column tubing not seated properly – creates a void Resulting peak shape Unswept volume causing brand broadening Proper Connection Resulting peak shape Tubing flush with end-fitting and good seal Importance of Batch-to-Batch and Column-to-Column Reproducibility ■■Column reproducibility is a key attribute...
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PART 2: BIOSEPARATION METHOD DEVELOPMENT Eluent Effect on SEC Peptide Separations ■■Non-desired, secondary interactions can compromise LC separations ■■Use of an appropriate LC eluent can minimize secondary interactions 25 mM sodium phosphate, pH 6.2, 250 mM sodium chloride Method development experiments evaluated the effect of mobile-phase pH and salt concentration. The results showed minimal effect of salt concentrations (150–350 mM) and mobile-phase pH (6.2–7.4) on retention time (data not shown). All of the aqueous mobile phases resulted in later than expected elution for most small peptides...
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Temperature Effect on Reversed-Phase Protein Separations ■■Use of “room temperature” is NOT always the ideal separation temperature ■■Use of a column heater is strongly recommended for reproducible analyses The intact IgG sample gave no observable peak at 40 °C, but recovery for the IgG sample improves with increasing temperature. There is not a measurable increase in recovery or improvement in peak shape above 80 °C. Column temperature has a large effect on reversedphase separation of molecules. Changes in recovery and selectivity are not uncommon with small molecule separations. While increasing...
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Effect of Gradient Duration of a Reversed-Phase Peptide Separation ■■Use of longer gradient time can translate into improved component resolution ■■Analysis time will increase using longer gradients as will peak volume ■■Sample complexity can influence selected gradient duration The use of salt gradients can also be used to analyze variants of a single protein. In this example, chicken albumin was analyzed by anion-exchange chromatography. Three different gradient slopes were employed to analyze the variants of albumin formed by post transitional modifications, such as methylation, phosphorylation,...
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[ WATERS ONLINE ORDER CENTER ] Whether you are a lab manager, scientist, or purchasing agent in procurement, we make it easy for you to research, compare, and purchase consumables and spare parts online.
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Amino Acid Analysis Amino Acid Analysis 9
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Amino Acid Analysis
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Amino Acid Analysis Amino acids are the constituents of proteins and are the intermediates in many metabolic pathways. Qualitative and quantitative Amino Acid Analysis (AAA) is used to determine the concentration of proteins, identify proteins, and detect structural variants. Amino acid composition is a critical component of the nutritional value of foods and feeds. The same analytical tools are used to monitor cell culture and fermentation processes. AAA is also used as a clinical diagnostic tool for assessing inborn errors of metabolism and nutritional status. The accurate identification and...
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ACCURATE AMINO ACID ANALYSES FROM VARIED SAMPLE MATRICES The UPLC Amino Acid Analysis Solution includes two complete methods using the same instrumentation and chemistries. The first is suitable for the amino acids derived from protein hydrolysates. The second is suitable for the larger number of free amino acids found in process samples such as cell culture or fermentation broths. The methods differ in the dilution of the AccQ•Tag Ultra Eluent A and the separation column temperature. There are no user adjustments of pH or modifications of composition for either Eluent A or Eluent B. Conditions...
Open the catalog to page 14Archived catalogs
XBridge™12 Pages
Torus™7 Pages
Enzymate BEH Pepsin Columns5 Pages
Viridis®6 Pages
Protein-Pak™6 Pages
Trefoil™7 Pages
ACQUITY UPLC M-Class3 Pages
Preparative OBD Columns Brochure12 Pages
24322 Pages
ACQUITY UPLC I-Class PLUS6 Pages
ACQUITY UPLC H-Class PLUS6 Pages
ProteinWorks6 Pages
PoraPak Rxn Brochure8 Pages
HPLC Column Performance30 Pages
Food Testing Solutions Brochure14 Pages
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