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Polymemse chain reaction using mixed primers. Cloning of human monoclonal antibody variable region genes from single hybridoma cells

Author:
  • James W Larrick
  • Lena Danielsson
  • Carol A Brenner
  • Ellen F Wallace
  • Magnus Abrahamson
  • Kirk E Fry
  • Carl Borrebaeck
Publishing year: 1989
Language: English
Pages: 934-938
Publication/Series: Nature Biotechnology
Volume: 7
Issue: 9
Document type: Journal article
Publisher: Nature Publishing Group

Abstract english

We describe a general approach to rapidly obtain the DNA sequence encoding the variable region of any immunoglobulin chain using the polymerase chain reaction and a mixture of upstream primers corresponding to the leader sequence, and one downstream primer designed from the conserved nucleotide sequence of the constant region. The approach was applied to five different hybridomas producing human monoclonal antibodies and variable regions for both bold gamma and mu heavy chain and kappa and lambda light chain genes were successfully cloned. cDNA encoding variable regions could be amplified from single hybridoma cells isolated by micromanipulation. This approach will permit analysis of B cell clonal ontogeny, antibody diversity and lymphoma cell progression and heterogeneity. It will also facilitate structural and functional studies of immunoglobulins as well as the rapid construction of chimeric antibodies.

Keywords

  • Pharmacology and Toxicology
  • Medicinal Chemistry

Other

Published
  • ISSN: 1546-1696
Carl B
Carl Borrebaeck
E-mail: carl [dot] borrebaeck [at] immun [dot] lth [dot] se

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Department of Immunotechnology

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