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Nat Methods. 2018 May;15(5):330-338. doi: 10.1038/nmeth.4632. Epub 2018 Mar 19.

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

Author information

1
Solomon H. Snyder Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
2
Department of Biomedical Engineering, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
3
CDI Laboratories, Mayaguez, PR, USA.
4
HudsonAlpha Institute for Biotechnology, Huntsville, Alabama, USA.
5
Institute for System Genetics, NYU Langone Health, New York, New York, USA.
6
Department of Biochemistry and Molecular Pharmacology, NYU Langone Health, New York, New York, USA.
7
Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
8
Department of Pharmacology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
9
Cancer Research Technology Program, Frederick National Laboratory for Cancer Research, Leidos Biomedical Research, Inc., Frederick, Maryland, USA.
10
Department of Psychiatry and Behavioral Sciences, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
11
Eukaryotic Tissue Culture Facility, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
12
NeoBiotechnologies, Inc., Union City, California, USA.
13
Department of Molecular Biology and Biochemistry, Rutgers University, Piscataway, New Jersey, USA.
14
Center for Advanced Biotechnology and Medicine, Piscataway, New Jersey, USA.
15
Center for Human Systems Biology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
16
Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
17
Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
18
Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.

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.

PMID:
29638227
PMCID:
PMC6063793
[Available on 2019-05-01]
DOI:
10.1038/nmeth.4632

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