Absolute Quantification of a Therapeutic Domain Antibody Using Ultra-Performance Liquid Chromatography–Mass Spectrometry and Immunoassay

Szapacs, Matthew; Urbanski, J J; Kehler, Jonathan; Wilson, Robert; Boram, Sharon L; Hottenstein, Charles; Citerone, David R.

doi:10.4155/bio.10.70

Cited by 26 publications

(10 citation statements)

References 17 publications

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“…There is no need to always have both LBA and LC-MS assays developed in parallel even though bioanalysts tend to do that during the initial stages of method development. It has been reported that the results generated by both types of assays have good agreement with each other in most cases from a statistical point of view [39,50,52]. In some cases, however, due to the complexity and diversity of emerging biologics drug portfolios, as discussed before, it is becoming a necessity to have both sets of data from LBA and LC-MS assays to provide complementary information to aid better decision making.…”

Section: Cases When Both Lba and Lc-ms Assay Are Recommendedmentioning

confidence: 88%

The Integration of Ligand Binding and LC-MS-Based Assays Into Bioanalytical Strategies for Protein Analysis

2014

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Both LBAs and LC-MS-based assays are reviewed and summarized for applications in quantitative protein analysis. A strategy for platform selection is proposed based on several factors that contribute to the complexities of bioanalysis of biologics. Protein types, multiple co-existing forms, post-translational modifications, and affinities to ADA, targets, and endogenous proteins need to be considered when selecting the most appropriate platform. Other factors, such as intended use of data, assay sensitivity, available reagents, and multiple analytes also impact on the choice of bioanalytical platform. At BMS, strategies for the seamless integration of both platforms are being implemented to provide not only PK/PD data of the molecules but also useful information of the amino acid structure and functional relationship of the proteins.

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Section: Cases When Both Lba and Lc-ms Assay Are Recommendedmentioning

confidence: 88%

The Integration of Ligand Binding and LC-MS-Based Assays Into Bioanalytical Strategies for Protein Analysis

2014

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“…Historically, this technique has proven to be robust for use during the quantitation of drugs and can be automated for sample preparation in various formats (e.g., 96well plate or online). In a recent example, SPE was successfully used as a sample prepara tion approach for the quantitation of a 13 kDa domain antibody, providing a detection limit in the low ng/ml when combined with short LC-MS/MS ana lysis [18]. In this particular example, SPE was successfully employed in a whole protein LC-MS/MS workflow, as well as an alternative approach involving the SPEbased cleanup of a tryptic digest of the same domain antibody.…”

Section: Sample Preparation and Analyte Purificationmentioning

confidence: 95%

Peptide and Protein Drug Analysis by MS: Challenges and Opportunities for The Discovery Environment

Campbell¹,

Blanc²

2011

Bioanalysis

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Straightforward assay development using MS has become commonplace in most modern pharmaceutical laboratories. In particular, MS is an invaluable tool in the discovery environment of this industry, making it possible to characterize the structures of target drugs and to screen large numbers of potential drug candidates in metabolism and pharmacokinetics studies, and much more. Furthermore, as drug portfolios expand to include biotherapeutic species, such as peptides and proteins, MS is there to meet any analytical challenges. In this article, general aspects of MS in the discovery environment are discussed, as well as what the future might hold.

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“…For instance, Neubert et al () developed a magnetic bead‐based immunoprecipitation method to quantify an anti‐drug antibody (ADA) with the existence of therapeutic proteins in monkey and human plasma, and considerably facilitated the immunogenicity investigation of an mAb. The parallel use of LBA and LC‐MS can also reveal specific pharmaceutical information such as biological activity and molecular integrity of biotherapeutics (Szapacs, Urbanski, & Kehler, ). For example, LBA and LC/MS have been applied to determine separately the concentrations of mAbs and unconjugated drugs in ADC bioanalysis (Kaur, ).…”

Section: Quantification Of Biotherapeutics In Biomatricesmentioning

confidence: 99%

Qualitative and quantitative characterization of protein biotherapeutics with liquid chromatography mass spectrometry

Shen

et al. 2016

Mass Spectrometry Reviews

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In the last decade, the advancement of liquid chromatography mass spectrometry (LC/MS) techniques has enabled their broad application in protein characterization, both quantitatively and qualitatively. Owing to certain important merits of LC/MS techniques (e.g., high selectivity, flexibility, and rapid method development), LC/MS assays are often deemed as preferable alternatives to conventional methods (e.g., ligand-binding assays) for the analysis of protein biotherapeutics. At the discovery and development stages, LC/MS is generally employed for two purposes absolute quantification of protein biotherapeutics in biological samples and qualitative characterization of proteins. For absolute quantification of a target protein in bio-matrices, recent work has led to improvements in the efficiency of LC/MS method development, sample treatment, enrichment and digestion, and high-performance low-flow-LC separation. These advances have enhanced analytical sensitivity, specificity, and robustness. As to qualitative analysis, a range of techniques have been developed to characterize intramolecular disulfide bonds, glycosylation, charge variants, primary sequence heterogeneity, and the drug-to-antibody ratio of antibody drug conjugate (ADC), which has enabled a refined ability to assess product quality. In this review, we will focus on the discussion of technical challenges and strategies of LC/MS-based quantification and characterization of biotherapeutics, with the emphasis on the analysis of antibody-based biotherapeutics such as monoclonal antibodies (mAbs) and ADCs. © 2016 Wiley Periodicals, Inc. Mass Spec Rev 36:734-754, 2017.

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Absolute Quantification of a Therapeutic Domain Antibody Using Ultra-Performance Liquid Chromatography–Mass Spectrometry and Immunoassay

Abstract: The two assays show good correlation (r(2) = 0.92), giving confidence in using either method for quantification of the dAb.

Cited by 26 publications

References 17 publications

The Integration of Ligand Binding and LC-MS-Based Assays Into Bioanalytical Strategies for Protein Analysis

The Integration of Ligand Binding and LC-MS-Based Assays Into Bioanalytical Strategies for Protein Analysis

Peptide and Protein Drug Analysis by MS: Challenges and Opportunities for The Discovery Environment

Qualitative and quantitative characterization of protein biotherapeutics with liquid chromatography mass spectrometry

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