重组Anti-Histone H2B抗体[mAbcam 52484] - ChIP Grade (ab52484)

Chromatin was prepared from HeLa cells according to the Abcam X-ChIP protocol*. Cells were fixed with formaldehyde for 10min.

The ChIP was performed with 25 µg of chromatin, 2 µg of ab52484 (red), or 2 µg of mouse IgG1 ab18443 (gray) and 25 µl of Protein A/G Dynabeads. The immunoprecipitated DNA was quantified by real time PCR (Sybr green approach).

NIH/3T3 cells were fixed in 100% methanol and permeabilized with 0.1% Triton X-100. Primary antibody, ab52484 at 1:2000 was incubated overnight at 4° C, followed by AlexaFluor® 488-conjugated Goat anti-mouse secondary antibody (ab150117) at 1/1000 dilution at RT for 45 min. ab179513 Anti-beta Tubulin, used as a counterstain at 1/200 dilution, was co-incubated with ab52484 overnight at 4° C, followed by Alexa Fluor® 594 Goat Anti-Rabbit secondary (ab150080) at 1/1000 dilution at RT for 45 min. Nucleus were visualized using DAPI.

HeLa cells were fixed in 100% methanol and permeabilized with 0.1% Triton X-100. Primary antibody, ab52484 at 1:2000 was incubated overnight at 4° C, followed by AlexaFluor® 488-conjugated Goat anti-mouse secondary antibody (ab150117) at 1/1000 dilution at RT for 45 min. ab179513 Anti-beta Tubulin, used as a counterstain at 1/200 dilution, was co-incubated with ab52484 overnight at 4° C, followed by Alexa Fluor® 594 Goat Anti-Rabbit secondary (ab150080) at 1/1000 dilution at RT for 45 min. Nucleus were visualized using DAPI.

Nuclear staining in rat lung. The section was heat mediated antigen retrieval with Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins. The section was incubated with ab52484 (1:50000 (0.021 μg/ml)) for 10 mins at room temperature, followed by anti-mouse IgG1 antibody (ab125913) for 8 mins during the LeicaDS9800 kit staining procedure.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Nuclear staining in mouse lung. The section was heat mediated antigen retrieval with Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins. The section was incubated with ab52484 (1:50000 (0.021 μg/ml)) for 10 mins at room temperature, followed by anti-mouse IgG1 antibody (ab125913) for 8 mins during the LeicaDS9800 kit staining procedure.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Nuclear staining in human breast cancer. The section was heat mediated antigen retrieval with Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins.

The section was incubated with ab52484 at 1:20000 (0.053 μg/ml) for 10 mins at room temperature, followed by anti-mouse IgG1 antibody (ab125913) for 8 mins during the LeicaDS9800 kit staining procedure.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Histone H2B was immunoprecipitated from 0.35 mg HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate with AB52484 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using AB52484 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution. 

Lane 1: HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate  10 ug

Lane 2: AB52484 IP in HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate

Lane 3: Mouse monoclonal IgG (ab18443) instead of AB52484 in HeLa whole cell lysate

Blocking and dilution buffer and concentration: 5% NFDM/TBST.

Exposure time: 32 seconds.

Flow cytometric analysis of NIH/3T3 (mouse embryonic fibroblast) cells labelling Histone H2B with AB52484 at 1/100 dilution (1 µg) (Red) compared with a Mouse monoclonal IgG (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). Goat anti mouse IgG (Alexa Fluor® 488, ab150113) at 1/2000 dilution was used as the secondary antibody.

Flow cytometric analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labelling Histone H2B with AB52484 at 1/1000 dilution (0.1µg) (Red) compared with a Mouse monoclonal IgG (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). Goat anti mouse IgG (Alexa Fluor® 488, ab150113) at 1/2000 dilution was used as the secondary antibody.

This image was generated using a previous batch manufactured using hybridoma production method.

ab52484 staining Histone H2B in Fruit fly (Drosophila melanogaster) embryo cells by ICC/IF (Immunocytochemistry/immunofluorescence). Embryos were washed in 1x PBS + 0.1% Trition, cells were fixed with heat, and blocked with 10% BSA for 12 hours at 4°C. Samples were incubated with primary antibody (1/1000) for 24 hours. An Alexa Fluor®594-conjugated Rabbit anti-mouse IgG polyclonal (1/5000) was used as the secondary antibody.

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This image was generated using a
previous batch manufactured using hybridoma production method