Toward low-cost affinity reagents: lyophilized yeast-scFv probes specific for pathogen antigens.

PloS One
Sean A GrayGerard A Cangelosi

Abstract

The generation of affinity reagents, usually monoclonal antibodies, remains a critical bottleneck in biomedical research and diagnostic test development. Recombinant antibody-like proteins such as scFv have yet to replace traditional monoclonal antibodies in antigen detection applications, in large part because of poor performance of scFv in solution. To address this limitation, we have developed assays that use whole yeast cells expressing scFv on their surfaces (yeast-scFv) in place of soluble purified scFv or traditional monoclonal antibodies. In this study, a nonimmune library of human scFv displayed on the surfaces of yeast cells was screened for clones that bind to recombinant cyst proteins of Entamoeba histolytica, an enteric pathogen of humans. Selected yeast-scFv clones were stabilized by lyophilization and used in detection assay formats in which the yeast-scFv served as solid support-bound monoclonal antibodies. Specific binding of antigen to the yeast-scFv was detected by staining with rabbit polyclonal antibodies. In flow cytometry-based assays, lyophilized yeast-scFv reagents retained full binding activity and specificity for their cognate antigens after 4 weeks of storage at room temperature in the absence of des...Continue Reading

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Citations

Jan 19, 2013·Chemical Communications : Chem Comm·Yadveer S GrewalMatt Trau
Nov 17, 2015·Journal of Immunological Methods·Maria KahnGonzalo J Domingo
Apr 12, 2015·Biosensors & Bioelectronics·A G VenkateshEliah Aronoff-Spencer
May 26, 2015·Clinics in Laboratory Medicine·Ibne Karim M Ali
Nov 5, 2016·ACS Applied Materials & Interfaces·Yadveer S GrewalMatt Trau

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Methods Mentioned

BETA
fluorescent-activated cell sorting
Immunoprecipitation
flow cytometry
FACS
light microscopy
ELISA
fluorescence microscopy
immuno-cytometry
Assay
PCR

Software Mentioned

ImageJ
Metamorph
Image J

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