HeLa cells were stained with the H3K9me1 antibody (bs-53043R) and with DAPI. Cells were formaldehyde fixated, permeabilized with Triton X-100 and blocked with PBS containing 2.5% BSA. Figure A: cells were immunofluorescently labelled with the H3K9me1 antibody (diluted 1:200 and incubated for 1 hour at room temperature) followed by goat anti-rabbit antibody conjugated to DyLight. Figure B: staining of the nuclei with DAPI, which specifically labels DNA. Both antibody and DAPI staining are restricted to the nucleus.


To determine the titer, an ELISA was performed using a serial dilution of the H3K9me1 antibody (bs-53043R). The antigen used was a peptide containing the histone modification of interest. By plotting the absorbance against the antibody dilution the titer of the antibody was estimated to be 1:50,000.


A Dot Blot analysis was performed to test the cross reactivity of H3K9me1 antibody (bs-53043R) with peptides containing other modifications of histone H3. Other histone modifications include di- and trimethylation of the same lysine and mono-, di- and trimethylation of lysine 27, 36 and 79. One hundred to 0.2 pmol of peptide containing the respective histone modification were spotted on a membrane. The antibody was used at a dilution of 1:200,000. The figure shows a high specificity of the antibody for the modification of interest.


Histone extracts of HeLa cells (15 μg) were analysed by Western blot using the H3K9me1 antibody (bs-53043R) diluted 1:1000 in TBS-Tween containing 5% skimmed milk. The position of the protein of interest is indicated on the left; the marker (in kDa) is shown on the right.