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Sequencing and Affinity Determination of Antigen-Specific B Lymphocytes from Peripheral Blood


Here we describe methods for screening human blood to isolate peripheral blood mononuclear cells (PBMCs) capable of binding fluorescently labeled antigen, as well as methods for the amplification and sequencing of B cell receptor (BCR) heavy and light chain genes. Detailed protocols are provided for transient mammalian expression in a hexahistidine-tagged Fab format, purification by immobilized metal affinity chromatography (IMAC), and affinity determination by BioLayer interferometry (BLI).

Type Book section
ISBN 1940-6029 (Electronic) 1064-3745 (Linking)
Authors Schofield, P.; Vazquez-Lombardi, R.; Abdelatti, M.; Nevoltris, D.; Goodnow, C. C.; Christ, D.; Reed, J. H.
Responsible Garvan Author Dr Joanne Reed
Publisher Name Methods in Molecular Biology
Published Date 2018-10-01
Published Volume 1827
Published Pages 287-309
Status Published in-print
DOI 10.1007/978-1-4939-8648-4_15
URL link to publisher's version