Refined protocol for generating monoclonal antibodies from single human and murine B cells

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• 作者：Irvin Y. Ho^a ; ¹ ; ^{irvin@uchicago.edu" class="auth_mail" title="E-mail the corresponding author} ; Jeffrey J. Bunker^b ; ^c ; ¹ ; ^{jeffbunker@uchicago.edu" class="auth_mail" title="E-mail the corresponding author} ; Steven A. Erickson^b ; ^c ; ^{sae@uchicago.edu" class="auth_mail" title="E-mail the corresponding author} ; Karlynn E. Neu^a ; ^c ; ^{karlynn@uchicago.edu" class="auth_mail" title="E-mail the corresponding author} ; Min Huang^a ; ^{huang2@uchicago.edu" class="auth_mail" title="E-mail the corresponding author} ; Mario Cortese^d ; ^{mario.cortese@emory.edu" class="auth_mail" title="E-mail the corresponding author} ; Bali Pulendran^d ; ^e ; ^{bpulend@emory.edu" class="auth_mail" title="E-mail the corresponding author} ; Patrick C. Wilson^a ; ^c ; ^{wilsonp@uchicago.edu" class="auth_mail" title="E-mail the corresponding author}
• 关键词：Monoclonal antibodies ; Single-cell PCR ; Human antibodies ; Gibson assembly ; Antibody expression
• 刊名：Journal of Immunological Methods
• 出版年：2016
• 出版时间：November 2016
• 年：2016
• 卷：438
• 期：Complete
• 页码：67-70
• 全文大小：377 K

文摘

Generating monoclonal antibodies from single B cells is a valuable tool for characterizing the specificity and functional properties of humoral responses. We and others developed protocols that have facilitated major advances in our understanding of B cell development, tolerance, and effector responses to HIV and influenza. Here, we demonstrate various refinements and dramatically reduce the time required to produce recombinant antibodies. Further, we present new methods for cloning and isolating antibodies from cells with lower immunoglobulin mRNA levels that may be resistant to traditional techniques. Together, these refinements significantly increase single-cell antibody expression efficiency and are easily integrated into established and novel pipelines.