CRISPR/Cas9 C-terminal Antibody - ChIP Grade (sample size)

Figure 1. ChIP using the Diagenode monoclonal antibody directed against Cas9
ChIP was performed on NIH3T3 cells stably expressing GFP-H2B, nuclease dead Cas9, and a GFP-targeting gRNA. 50 μg chromatin was incubated overnight at 4°C with 5 μg of either the Diagenode antibody against Cas9 (cat. No. C15200231) or the anti-blue antibody (C15200213), used as a negative IP control. qPCR was performed with primers speci c for the GFP gene, and for a non-targeted region (protein kinase C delta, Prkcd), used as negative control. Figure 1 shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).

Figure 2. Western blot analysis using the Diagenode monoclonal antibody directed against CRISPR/Cas9
Western blot was performed on protein extracts from HeLa cells transfected with Cas9 (lane 2) or on untransfected control cells (lane 1) using the Diagenode antibody against CRISPR/Cas9 (Cat. No. C15200231). The antibody was diluted 1:6,000 in TBS-T containing 3% NFDM. The marker is shown on the left, position of the Cas9 protein is indicated on the right.

Figure 3. Immunofluorescence using the Diagenode monoclonal antibody directed against CRISPR/Cas9
HeLa cells expressing Cas9 under the control of the tight TRE promoter and untransfected control cells were fixed in methanol at -20°C, permeabilized with acetone at -20°C and blocked with PBS containing 2% BSA. The cells were stained with the Cas9 C-terminal antibody (Cat. No. C15200231) diluted 1:100, followed by incubation with a donkey anti-mouse secondary antibody coupled to AF488 (upper panel). Nuclei were counter-stained with Hoechst 33342 (lower panel).