Histone H3 [p Thr11] Antibody

Western Blot: Histone H3 [p Thr11] Antibody [NBP2-59274]

Western Blot: Histone H3 [p Thr11] Antibody [NBP2-59274] - Western blot was performed on whole cell extracts from untreated HeLa cells (25 ug, lane 1), on histone extracts from HeLa cells treated with colcemid (15 ug, lane 2), and on 1 ug of recombinant histone H2A, H2B, H3 and H4 (lane 3, 4, 5 and 6, respectively) using the antibody against H3T11p . The antibody was diluted 1:1,000 in TBS-Tween containing 5% skimmed milk. Observed molecular weight is ~15 kDa.

Chromatin Immunoprecipitation: Histone H3 [p Thr11] Antibody [NBP2-59274] - ChIP assays were performed using human HeLa cells, treated with colcemid, the antibody against H3T11p and optimized PCR primer sets for qPCR. ChIP was performed using sheared chromatin from 1.5 million cells. A titration of the antibody consisting of 0.5, 1, 2 and 5 ug per ChIP experiment was analysed. IgG (2 ug/IP) was used as negative IP control. QPCR was performed with primers for the EIF4A2 and GAPDH promoters, used as positive controls, and for the coding region of the inactive MYT1 gene and the Sat2 satellite repeat, used as negative controls. Figure shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).

Dot Blot: Histone H3 [p Thr11] Antibody [NBP2-59274] - To test the cross reactivity of the antibody against H3T11p, a Dot Blot analysis was performed with peptides containing other histone phosphorylations and the unmodified H3T11. One hundred to 0.2 pmol of the respective peptides were spotted on a membrane. The antibody was used at a dilution of 1:20,000. Figure shows a high specificity of the antibody for the modification of interest.