近日，Wan CC等科学家在Nature Biotechnology上介绍了一种采用蛋白质组学法鉴定免疫动物血清中抗原特异性抗体的技术。这项技术采用了包括高特异性的抗体亲和纯化、nano-flow LC/MS分析消化的抗体片段，以及从已有的数据库中比对抗体分子高变区等。最终，重链和轻链都表达于重组单体。通过这个技术，该小组从小鼠和兔子的血清中纯化了5种了抗原特异性抗体，并发现这些单抗的活性超过了他们原始的多抗。可以预见，这项技术可以揭示和发展疫苗及抗体战略，以及帮助我们更深入的理解体液免疫。

 

 

A proteomics approach for the identification and cloning of monoclonal antibodies from serum

 

We describe a proteomics approach that identifies antigen-specific antibody sequences directly from circulating polyclonal antibodies in the serum of an immunized animal. The approach involves affinity purification of antibodies with high specific activity and then analyzing digested antibody fractions by nano-flow liquid chromatography coupled to tandem mass spectrometry. High-confidence peptide spectral matches of antibody variable regions are obtained by searching a reference database created by next-generation DNA sequencing of the B-cell immunoglobulin repertoire of the immunized animal. Finally, heavy and light chain sequences are paired and expressed as recombinant monoclonal antibodies. Using this technology, we isolated monoclonal antibodies for five antigens from the sera of immunized rabbits and mice. The antigen-specific activities of the monoclonal antibodies recapitulate or surpass those of the original affinity-purified polyclonal antibodies. This technology may aid the discovery and development of vaccines and antibody therapeutics, and help us gain a deeper understanding of the humoral response.

 

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