The authors describe how for every chromatin preparation it is essential to check the chromatin is sheared to fragments between 100-500 bp. They emphasise using a microfluidics platform is the most accurate measure of quantifying DNA.
Problems associated with over or under sheared Chromatin fragments are discussed and a protocol for optimised enzymatic shearing described. As some cells are resistant to lysis the authors also describe an optimised sonication methodology to ensure a successful ChIP result.
Launched worldwide in 2012 - Chromatrap® solid-state ChIP technology has been shown by a growing number of research groups worldwide to be more efficient than conventional bead-based methods. This is because the solid phase porous polymer, functionalized with either protein A or G, provides a greater surface area for chromatin antibody binding with very low non-specific binding. In addition, it uses a spin column approach, offering significant speed, process and carry-over advantages over sepharose or magnetic beads. DNA pull down with Chromatrap® is up to 25 times more than conventional methods, whilst the signal to noise ratio for DNA enrichment is 2 to 3 times better, even with low chromatin samples between 50ng to 3000ng per immunoprecipitation.
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