For over 15 years Diagenode has been the most trusted supplier for epigenetics-focused antibodies. We understand that high quality is crucial for the success of experiments, so we emphasize strict quality standards and the actual validation for all of our antibodies in applications such ChIP and ChIP-seq. Our antibodies are produced and validated following the ChIP-seq guidelines defined by major epigenetic consortia including Blueprint and ENCODE.
New interest is emerging for new applications for antibody-targeted chromatin profiling: CUT&RUN (Cleavage Under Targets and Release Using Nuclease) and CUT&Tag (Cleavage Under Targets and Tagmentation). In CUT&RUN, the protein of interest is bound by a specific antibody, the protein A-Mnase fusion binds to an antibody targeting the protein of interest, and Mnase cleaves adjacent DNA upon calcium activation. DNA bound to the protein of interest is released and extracted, after which libraries for sequencing can be prepared. The second method, CUT&Tag uses protein A-Tn5 fusion (preloaded with sequencing adaptors) instead of protein A-Mnase. Protein A-Tn5 complex binds to the antibody of interest and upon the activation with magnesium, the Tn5 transposase cuts adjacent DNA and inserts sequencing adaptors, generating libraries ready for sequencing.
The quality of antibody used in CUT&RUN or in CUT&Tag is one of the crucial factors for assay success. The antibodies with confirmed high specificity will target only the protein of interest, enabling real results. All Diagenode ChIP-seq grade antibodies are validated for specificity using strict validation criteria to assure high performance in CUT&Tag.
Figure 1. Representative screenshot for H3K4me3 (A and B), H3K27me3 (C and D), H3K9me3 (E and F) data obtained using CUT&Tag protocol (Kaya-Okur, H.S., Nat Commun 10, 1930 (2019)) at selected loci. CUT&Tag was performed using 50,000 K562 cells and Diagenode H3K4me3 polyclonal ChIP-seq antibody (Cat. No. C15410003), H3K27me3 polyclonal ChIP-seq grade antibody (Cat. No. C15410195) and H3K9me3 poyclonal ChIP-seq grade antibody (Cat. No. C15410193).
Figure 2. Typical library profile generated by CUT&Tag protocol. Agilent BioAnalyzer trace of pooled libraries derived from H3K4me3, H3K27me3 and H3K9me3 CUT&Tag experiments performed using 50,000 K562 cells (A) and from H3K27me3 and H3K9me3 CUT&Tag experiments using 25 000, 10 000 and 1 000 K562 cells (B). The following Diagenode antibodies were used: H3K4me3 polyclonal ChIP-seq antibody (Cat. No. C15410003), H3K27me3 polyclonal ChIP-seq grade antibody (Cat. No. C15410195) and H3K9me3 polyclonal ChIP-seq grade antibody (Cat. No. C15410193).