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Figure 1. ChIP results obtained with the Diagenode antibody directed against YY1 ChIP assays were performed using K562 cells, the Diagenode antibody against YY1 (Cat. No. C15410345) and optimized PCR primer sets for qPCR. ChIP was performed with the “iDeal ChIP-seq” kit (Cat. No. C01010055), using sheared chromatin from 4 million cells. A titration consisting of 1, 2, 5 and 10 μg of antibody per ChIP experiment was analyzed. IgG (2 μg/IP) was used as a negative IP control. Quantitative PCR was performed with primers for the promoters of the FBXL18 and EIF2S3 genes, used as positive controls, and for the 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. ChIP-seq results obtained with the Diagenode antibody directed against YY1 ChIP was performed with 1 μg of the Diagenode antibody against YY1 (Cat. No. C15410345) on sheared chromatin from 4,000,000 K562 cells using the “iDeal ChIP-seq” kit as described above. The IP’d DNA was subsequently analysed on an Illumina HiSeq 4000. Library preparation, cluster generation and sequencing were performed according to the manufacturer’s instructions. The 50 bp tags were aligned to the human genome using the BWA algorithm. Figure 2 shows the signal distribution along the complete sequence and a 1 Mb region of the human X-chromosome (figures 2A and B), and in two genomic regions surrounding the FBXL18 and EIF2S3 positive control genes (figure 2C and D).
Figure 3. Determination of the antibody titer To determine the titer of the antibody, an ELISA was performed using a serial dilution of Diagenode antibody directed against YY1 (Cat. No. C15410345). The plates were coated with the peptides used for immunization of the rabbit. By plotting the absorbance against the antibody dilution (Figure 3), the titer of the antibody was estimated to be >1:1,000,000.
Figure 4. Western blot analysis using the Diagenode antibody directed against YY1 Whole cell extracts from K562 cells were analysed by Western blot using the Diagenode antibody against YY1 (Cat. No. C15410345) diluted 1:1,000 in TBS-Tween containing 5% skimmed milk. The position of the protein of interest is indicated on the right; the marker (in kDa) is shown on the left.
WB Western blot : The quality of antibodies used in this technique is crucial for correct and specific protein identification. Diagenode offers huge selection of highly sensitive and specific western blot-validated antibodies.
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