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In vitro generation of anti-hepatitis B monoclonal antibodies from a single plasma cell using single-cell RT-PCR and cell-free protein synthesis

Paper ID Volume ID Publish Year Pages File Format Full-Text
21546 43227 2010 8 PDF Available
Title
In vitro generation of anti-hepatitis B monoclonal antibodies from a single plasma cell using single-cell RT-PCR and cell-free protein synthesis
Abstract

Monoclonal antibodies (mAbs) are an effective tool in therapeutics and diagnostics. A novel approach called the single-cell RT-PCR-linked in vitro expression system (SICREX) enables the high-throughput generation and screening of mAbs from single B cells. In this paper, instead of using B cells, cDNAs were synthesized from single plasma cells of an immunized mouse spleen. The light chain (Lc) and the Fd portion of the heavy chain (Hc) genes of each cell were amplified separately and followed by overlapping PCR to add a T7 promoter, a ribosome-binding site, and a T7 terminator. The paired Lc and Hc genes were simultaneously expressed by an Escherichia coli in vitro transcription and translation system followed by ELISA to measure their affinity for the antigen. A Fab fragment with affinity against the antigen was obtained from plasma cells of an immunized mouse with hepatitis B surface antigen (HBsAg).

Keywords
Single-cell; RT-PCR; Cell-free protein synthesis; Monoclonal antibodies; Hepatitis B
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In vitro generation of anti-hepatitis B monoclonal antibodies from a single plasma cell using single-cell RT-PCR and cell-free protein synthesis
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Bioscience and Bioengineering - Volume 109, Issue 1, January 2010, Pages 75–82
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us