MicroChIP--a rapid micro chromatin immunoprecipitation assay for small cell samples and biopsies.
Chromatin immunoprecipitation (ChIP) is a powerful technique for studying protein-DNA interactions. Drawbacks of current ChIP assays however are a requirement for large cell numbers, which limits applicability of ChIP to rare cell samples, and/or lengthy procedures with limited applications. There are to date no protocols for fast and parallel ChIPs of post-translationally modified histones from small cell numbers or biopsies, and importantly, no protocol allowing for investigations of transcription factor binding in small cell numbers. We report here the development of a micro (micro) ChIP assay suitable for up to nine parallel quantitative ChIPs of modified histones or RNA polymerase II from a single batch of 1000 cells. MicroChIP can also be downscaled to monitor the association of one protein with multiple genomic sites in as few as 100 cells. MicroChIP is applicable to small fresh tissue biopsies, and a cross-link-while-thawing procedure makes the assay suitable for frozen biopsies. Using MicroChIP, we characterize transcriptionally permissive and repressive histone H3 modifications on developmentally regulated promoters in human embryonal carcinoma cells and in osteosarcoma biopsies. muChIP creates possibilities for multiple parallel and rapid transcription factor binding and epigenetic analyses of rare cell and tissue samples.
Dahl
John Arne
JA
Institute of Basic Medical Sciences, Department of Biochemistry, Faculty of Medicine, University of Oslo, 0317 Oslo, Norway.
Collas
Philippe
P
D001665:Binding Sites; D001706:Biopsy; D001859:Bone Neoplasms / Q000235:genetics / Q000473:pathology; D018236:Carcinoma, Embryonal / Q000235:genetics / Q000473:pathology; D002648:Child; D047369:Chromatin Immunoprecipitation / Q000379:methods*; D006657:Histones / Q000032:analysis* / Q000378:metabolism; D006801:Humans; D008297:Male; D012516:Osteosarcoma / Q000235:genetics / Q000473:pathology; D011401:Promoter Regions, Genetic; D012319:RNA Polymerase II / Q000032:analysis / Q000378:metabolism; D013997:Time Factors; D014157:Transcription Factors / Q000032:analysis* / Q000378:metabolism
D006657:Histones; D014157:Transcription Factors; D012319:RNA Polymerase II