Automated Screening of Monoclonal Antibodies for SISCAPA Assays Using a Magnetic Bead Processor and Liquid Chromatography-selected Reaction Monitoring-mass Spectrometry

Overview

Journal J Immunol Methods

Publisher Elsevier

Specialties Allergy & Immunology
Pathology

Date 2009 Dec 8

PMID 19961853

Citations 29

Authors

Regine M Schoenherr

Lei Zhao

Jeffrey R Whiteaker

Li-Chia Feng

Li Li

Lina Liu

Xiuwen Liu

Amanda G Paulovich

Affiliations

Soon will be listed here.

Abstract

Stable Isotope Standards and Capture by Anti-Peptide Antibodies (SISCAPA) utilizes antibodies to enrich peptides from complex matrices for quantitation by stable isotope dilution mass spectrometry. SISCAPA improves sensitivity and limits the sample handling required for plasma-based analysis. Thus far, SISCAPA assays have been performed using polyclonal antibodies, yet monoclonal antibodies are an attractive alternative since they provide exquisite specificity, a renewable resource, and the potential for isolation of clones with very high affinities (10(-9) M or better). The selection of a good monoclonal antibody out of hundreds-to-thousands of clones presents a challenge, since the screening assay should ideally be in the format of the final SISCAPA assay, but performing the assays manually is labor- and time-intensive. In this manuscript, we demonstrate that monoclonal antibodies can be used in SISCAPA assays, and we describe an automated high-throughput SISCAPA method that makes screening of large numbers of hybridomas feasible while conserving time and resources.

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