A toolbox of immunoprecipitation-grade monoclonal antibodies to human transcription factors

Anand Venkataraman ^1,iD, Kun Yang ², Jose Irizarry ³, Mark Mackiewicz ^4,iD, Paolo Mita ^5,6,7,iD, Zheng Kuang ^6,19,iD, Lin Xue ¹, Devlina Ghosh ¹, Shuang Liu ⁸, Pedro Ramos ³, Shaohui Hu ³, Diane Bayron Kain ^3,19, Sarah Keegan ^5,6, Richard Saul ⁸, Simona Colantonio ⁹, Hongyan Zhang ⁸, Florencia Pauli-Behn ⁴, Guang Song ⁸, Edisa Albino ³, Lillyann Asencio ³, Leonardo Ramos ³, Luvir Lugo ³, Gloriner Morell ³, Javier Rivera ³, Kimberly Ruiz ³, Ruth Almodovar ³, Luis Nazario ³, Keven Murphy ³, Ivan Vargas ³, Zully Ann Rivera-Pacheco ³, Christian Rosa ³, Moises Vargas ³, Jessica McDade ⁷, Brian S Clark ¹, Sooyeon Yoo ¹, Seva G Khambadkone ¹⁰, Jimmy de Melo ¹, Milanka Stevanovic ¹, Lizhi Jiang ¹, Yana Li ¹¹, Wendy Y Yap ³, Brittany Jones ¹², Atul Tandon ¹², Elliot Campbell ^13,14, Gaetano T Montelione ^13,14, Stephen Anderson ^13,14, Richard M Myers ⁴, Jef D Boeke ^5,6,7, David Fenyö ^5,6, Gordon Whiteley ⁹, Joel S Bader ², Ignacio Pino ^3,iD, Daniel J Eichinger ^3,iD, Heng Zhu ^8,15,iD, Seth Blackshaw

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Abstract

A key component of efforts to address the reproducibility crisis in biomedical research is the development of rigorously validated and renewable protein-affinity reagents. As part of the US National Institutes of Health (NIH) Protein Capture Reagents Program (PCRP), we have generated a collection of 1,406 highly validated immunoprecipitation- and/or immunoblotting-grade mouse monoclonal antibodies (mAbs) to 737 human transcription factors, using an integrated production and validation pipeline. We used HuProt human protein microarrays as a primary validation tool to identify mAbs with high specificity for their cognate targets. We further validated PCRP mAbs by means of multiple experimental applications, including immunoprecipitation, immunoblotting, chromatin immunoprecipitation followed by sequencing (ChIP-seq), and immunohistochemistry. We also conducted a meta-analysis that identified critical variables that contribute to the generation of high-quality mAbs. All validation data, protocols, and links to PCRP mAb suppliers are available at http://proteincapture.org.

Footnotes:
These authors contributed equally: Anand Venkataraman ^1,iD, Kun Yang ², Jose Irizarry ³, Mark Mackiewicz ^4,iD, Paolo Mita ^5,6,7,iD, Zheng Kuang ^6,19,iD, Lin Xue ¹, Devlina Ghosh ¹, Shuang Liu ⁸, Pedro Ramos ³, Shaohui Hu ³, Diane Bayron Kain ^3,19, Sarah Keegan ^5,6, Richard Saul ⁸, Simona Colantonio ⁹, Hongyan Zhang ⁸, Florencia Pauli-Behn ⁴, Guang Song ⁸, Edisa Albino ³, Lillyann Asencio ³, Leonardo Ramos ³, Luvir Lugo ³, Gloriner Morell ³, Javier Rivera ³, Kimberly Ruiz ³, Ruth Almodovar ³, Luis Nazario ³, Keven Murphy ³, Ivan Vargas ³, Zully Ann Rivera-Pacheco ³, Christian Rosa ³, Moises Vargas ³, Jessica McDade ⁷, Brian S Clark ¹, Sooyeon Yoo ¹, Seva G Khambadkone ¹⁰, Jimmy de Melo ¹, Milanka Stevanovic ¹, Lizhi Jiang ¹, Yana Li ¹¹, Wendy Y Yap ³, Brittany Jones ¹², Atul Tandon ¹², Elliot Campbell ^13,14, Gaetano T Montelione ^13,14, Stephen Anderson ^13,14, Richard M Myers ⁴, Jef D Boeke ^5,6,7, David Fenyö ^5,6, Gordon Whiteley ⁹, Joel S Bader ², Ignacio Pino ^3,iD, Daniel J Eichinger ^3,iD, Heng Zhu ^8,15,iD, Seth Blackshaw

Competing Interests: S.B., H. Zhu, I.P., D.J.E., and J.D.B. are cofounders and shareholders of CDI Labs Inc. J.I., P.R., D.B.K., E.A., L.A., L.R., L.L., G.M., J.R., K.R., R.A., L.N., K.M., I.V., Z.A.R.-P., C.R., M.V., and W.Y.Y. are employees of CDI Labs Inc. A.V. and J.D.B. are consultants to CDI Labs Inc. J.D.B. serves on the Board of Directors of CDI Labs, and J.D.B.’s relationship with CDI Labs is managed by NYU Langone Health’s committee on conflicts of interest. B.J. is an employee of NeoBiotechnologies, Inc. A.T. is the founder and sole owner of NeoBiotechnologies, Inc. G.T.M. is founder and shareholder of Nexomics Biosciences, Inc.
DOI: http://10.1038/nmeth.4632