WB
Western blot analysis of lysates from C6 cells using Phospho-Histone H3-S10 Rabbit pAb at 1:6000 dilution incubated overnight at 4℃. C6 cells were treated with nocodazole (100 nM) at 37°C for 24 hours.
Secondary antibody: HRP-conjugated Goat anti-Rabbit IgG (H+L) at 1:10000 dilution.
Lysates/proteins: 30 μg per lane.
Blocking buffer: 3% nonfat dry milk in TBST.
Detection: ECL Basic Kit.
Exposure time: 45 s.
DB
Dot-blot analysis of all sorts of peptides using Phospho-Histone H3-S10 Rabbit pAb at 1:1000 dilution.
IHC
Immunohistochemistry analysis of paraffin-embedded Mouse testis tissue, untreated (left) and lambda phosphatase-treated (right), tissue using Phospho-Histone H3-S10 Rabbit pAb at a dilution of 1:3000 (40x lens). High pressure antigen retrieval performed with 0.01M Tris-EDTA Buffer (pH 9.0) prior to IHC staining.
ICC/IF
Confocal imaging of HeLa cells (treated with nocodazole) and HeLa cells (untreated) using Phospho-Histone H3-S10 Rabbit pAb (dilution 1:2000) followed by a further incubation with Cy3-conjugated Goat anti-Rabbit IgG (H+L) (dilution 1:500) (Red). DAPI was used for nuclear staining (Blue). Objective: 100x.
CHIP
Chromatin immunoprecipitation was performed with 15 μg of cross-linked chromatin from HeLa cells were treated with nocodazole (100ng/ml 16h), using 3 μg of Phospho-Histone H3-S10 Rabbit pAb and Rabbit IgG isotype control. The enrichment of immunoprecipitated DNA at different genomic loci was examined by quantitative PCR. The histogram compares the ratio of the immunoprecipitated DNA to the input at given loci.