Capture and display of antibodies secreted by hybridoma cells enables fluorescent on-cell screening

MAbs
Rama Devudu PuligeddaScott K Dessain

Abstract

Hybridoma methods for monoclonal antibody (mAb) cloning are a mainstay of biomedical research, but they are hindered by the need to maintain hybridomas in oligoclonal pools during antibody screening. Here, we describe a system in which hybridomas specifically capture and display the mAbs they secrete: On-Cell mAb Screening (OCMS™). In OCMS™, mAbs displayed on the cell surface can be rapidly assayed for expression level and binding specificity using fluorescent antigens with high-content (image-based) methods or flow cytometry. OCMS™ demonstrated specific mAb binding to poliovirus and rabies virus by forming a cell surface IgG "cap", as a universal assay for anti-viral mAbs. We produced and characterized OCMS™-enabled hybridomas secreting mAbs that neutralize poliovirus and used fluorescence microscopy to identify and clone a human mAb specific for the human N-methyl-D-aspartate receptor. Lastly, we used OCMS™ to assess expression and antigen binding of a recombinant mAb produced in 293T cells. As a novel method to physically associate mAbs with the hybridomas that secrete them, OCMS™ overcomes a central challenge to hybridoma mAb screening and offers new paradigms for mAb discovery and production.

References

May 1, 1986·Journal of Immunological Methods·W L CarrollR Levy
Mar 14, 1995·Proceedings of the National Academy of Sciences of the United States of America·R ManzA Radbruch
Jun 1, 1997·Protein Science : a Publication of the Protein Society·S FreitagR E Stenkamp
Aug 1, 1995·Bio/technology·J S KenneyJ F Dunne
Dec 11, 1999·Nucleic Acids Research·H M BermanP E Bourne
Mar 5, 2004·Proceedings of the National Academy of Sciences of the United States of America·Keigo MachidaMichael M C Lai
Jul 21, 2004·Journal of Immunological Methods·Michael J Feldhaus, Robert W Siegel
Nov 1, 2005·Analytical Chemistry·Zhihong ShenXiangqun Zeng
Jun 26, 2009·Methods in Molecular Biology·Mitchell Ho, Ira Pastan
Apr 26, 2012·MAbs·Stacey SpencerYiqing Feng
Jul 4, 2012·Proceedings of the National Academy of Sciences of the United States of America·Bachir El DebsChristoph A Merten
Aug 10, 2012·The Journal of Neuroscience : the Official Journal of the Society for Neuroscience·Amy J GleichmanDavid R Lynch
Nov 28, 2013·Proceedings of the National Academy of Sciences of the United States of America·Zhaochun ChenKonstantin Chumakov
May 16, 2014·Antiviral Research·Rama Devudu PuligeddaKonstantin Chumakov
Jun 25, 2015·Microbiology Spectrum·Scott A Smith, James E Crowe
Mar 15, 2016·Expert Opinion on Drug Discovery·Ivan Fraietta, Fabio Gasparri
Apr 28, 2016·Trends in Cell Biology·Mojca Mattiazzi UsajBrenda J Andrews
Jun 9, 2016·Biotechnology Journal·Joseph J PriolaMichael J Betenbaugh
Mar 1, 2013·Biophysics Reviews·Daniel LaytonEdouard C Nice
Nov 22, 2017·Annals of Clinical and Translational Neurology·Manish MalviyaNorbert Goebels
Aug 22, 2018·Annals of Clinical and Translational Neurology·Rashmi SharmaScott K Dessain

❮ Previous
Next ❯

Methods Mentioned

BETA
fluorescence imaging
fluorescence microscopy
ELISA
flow cytometry
glycosylation
confocal microscopy
transfection
imaging techniques
ELISAs
fluorescence

Key Resources (RRID) Mentioned

AB2534088

Software Mentioned

ImageJ
GenScript

Related Concepts

Related Feeds

Antivirals

Antivirals are medications that are used specifically for treating viral infections. Discover the latest research on antivirals here.

Antivirals (ASM)

Antivirals are medications that are used specifically for treating viral infections. Discover the latest research on antivirals here.