H3K36me3 Antibody - ChIP Grade (sample size)

Figure 1. ChIP results obtained with the monoclonal antibody directed against H3K36me3
ChIP assays were performed using human HeLa cells, the monoclonal antibody against H3K36me3 (cat. No. C15200183), and optimized PCR primer pairs for qPCR. ChIP was performed using sheared chromatin from 1 million cells. A titration of 0.5, 1, 2, and 5 µg of antibody per ChIP experiment was analyzed. IgG (1 µg/IP) was used as a negative IP control. Quantitative PCR was performed with primers specific for the coding regions of the active EIF2S3 and CCT5 genes (used as positive controls) and for the inactive MYOD1 gene and the Sat2 satellite repeat (used as negative controls). 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. Cut&Tag results obtained with the monoclonal antibody directed against H3K36me3
Cut&Tag was performed on 50,000 K562 cells using 1 µg of the monoclonal antibody against H3K36me3 (cat. No. C15200183), the pA-Tn5 transposase (C01070001), and the iDeal Cut&Tag kit (cat. No. C01070021). The libraries were subsequently analyzed on an Illumina NovaSeq sequencer (2x50 paired-end reads) according to the manufacturer’s instructions. The tags were aligned to the human genome (hg38) using the BWA algorithm. Figure 2 shows the peak distribution along the complete sequence and a 2 Mb region of the X chromosome (Figure 2A and 2B), and in two genomic regions on chromosomes 7 and 12 (Figure 2C and 2D, respectively).

Figure 3. Cross reactivity of the monoclonal antibody directed against H3K36me3
To test the specificity, an ELISA was performed using a serial dilution of the monoclonal antibody against H3K36me3 (cat. No. C15200183). The wells were coated with peptides containing the unmodified H3K36 region as well as the mono-, di-, and trimethylated H3K36 and the trimethylated H3K9. Figure 3 shows a high specificity of the antibody for the peptide containing the modification of interest.

Figure 4.Immunofluorescence using the monoclonal antibody directed against H3K36me3
HeLa cells were stained with an antibody against H3K36me3 (cat. No. C15200183) and with DAPI. Cells were fixed with methanol and blocked with PBS/TX-100 containing 5% normal goat serum and 1% BSA. The cells were immunofluorescently labeled with the H3K36me3 antibody (left), diluted 1:500 in blocking solution, followed by an anti-mouse antibody conjugated to Alexa594. The middle panel shows staining of the nuclei with DAPI. A merge of the two stains is shown on the right.