Analysis of chromatin binding dynamics using the crosslinking kinetics (CLK) method

•A method for measuring chromatin-binding dynamics in vivo is described.•A quench flow apparatus is adapted to obtain kinetic data on the sub-second scale.•The protocol includes strategies for key decision points.•Data analysis and interpretation are discussed. Transcription factor binding sites in chromatin are routinely inventoried by the chromatin immunoprecipitation assay, and these binding patterns can provide precise and detailed information about cell state. However, some fundamental molecular questions regarding transcription factor function require an understanding of in vivo binding dynamics as well as location information. Here we describe the crosslinking kinetics (CLK) assay, in which the time-dependence of formaldehyde crosslinking is used to extract on- and off-rates for chromatin binding in vivo.