A toolbox of immunoprecipitation-grade monoclonal antibodies against human transcription factors.
Florencia Pauli Behn,
Zully Ann Rivera-Pacheco,
Brian S. Clark,
Seva G Khambadkone,
Jimmy de Melo,
Wendy Y Yap,
Richard M Myers,
Jef D. Boeke,
Joel S Bader,
Daniel J Eichinger,
Posted 14 Mar 2017
bioRxiv DOI: 10.1101/116442
Posted 14 Mar 2017
A key component to overcoming the reproducibility crisis in biomedical research is the development of readily available, rigorously validated and renewable protein affinity reagents. As part of the NIH Protein Capture Reagents Program (PCRP), we have generated a collection of 1406 highly validated, immunoprecipitation (IP) and/or immunoblotting (IB) grade, mouse monoclonal antibodies (mAbs) to 736 human transcription factors. We used HuProt human protein microarrays to identify mAbs that recognize their cognate targets with exceptional specificity. Using an integrated production and validation pipeline, we validated these mAbs in multiple experimental applications, and have distributed them to the Developmental Studies Hybridoma Bank (DSHB) and several commercial suppliers. This study allowed us to perform a meta-analysis which identified critical variables that contribute to the generation of high quality mAbs. We find that using full-length antigens for immunization, in combination with HuProt analysis, provides the highest overall success rates. The efficiencies built into this pipeline ensure substantial cost savings compared to current standard practices.
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