2016年4月
Novel method for the high-throughput production of phosphorylation site-specific monoclonal antibodies
SCIENTIFIC REPORTS
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- 巻
- 6
- 号
- 開始ページ
- 25174
- 終了ページ
- 記述言語
- 英語
- 掲載種別
- 研究論文(学術雑誌)
- DOI
- 10.1038/srep25174
- 出版者・発行元
- NATURE PUBLISHING GROUP
Threonine phosphorylation accounts for 10% of all phosphorylation sites compared with 0.05% for tyrosine and 90% for serine. Although monoclonal antibody generation for phospho-serine and -tyrosine proteins is progressing, there has been limited success regarding the production of monoclonal antibodies against phospho-threonine proteins. We developed a novel strategy for generating phosphorylation site-specific monoclonal antibodies by cloning immunoglobulin genes from single plasma cells that were fixed, intracellularly stained with fluorescently labeled peptides and sorted without causing RNA degradation. Our high-throughput fluorescence activated cell sorting-based strategy, which targets abundant intracellular immunoglobulin as a tag for fluorescently labeled antigens, greatly increases the sensitivity and specificity of antigen-specific plasma cell isolation, enabling the high-efficiency production of monoclonal antibodies with desired antigen specificity. This approach yielded yet-undescribed guinea pig monoclonal antibodies against threonine 18-phosphorylated p53 and threonine 68-phosphorylated CHK2 with high affinity and specificity. Our method has the potential to allow the generation of monoclonal antibodies against a variety of phosphorylated proteins.
- リンク情報
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- DOI
- https://doi.org/10.1038/srep25174
- J-GLOBAL
- https://jglobal.jst.go.jp/detail?JGLOBAL_ID=201702205469729198
- PubMed
- https://www.ncbi.nlm.nih.gov/pubmed/27125496
- Web of Science
- https://gateway.webofknowledge.com/gateway/Gateway.cgi?GWVersion=2&SrcAuth=JSTA_CEL&SrcApp=J_Gate_JST&DestLinkType=FullRecord&KeyUT=WOS:000375179200001&DestApp=WOS_CPL
- ID情報
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- DOI : 10.1038/srep25174
- ISSN : 2045-2322
- J-Global ID : 201702205469729198
- PubMed ID : 27125496
- Web of Science ID : WOS:000375179200001