Anti-Histone H3 (phospho S28) antibody [HTA28]

• ICC/IF

AbReview40766****

Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (phospho S28) antibody [HTA28] (AB10543)

ab10543 staining Histone H3 (phospho S28) in Human neural progenitor cells from iPS cell line by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde, permeabilized with 0.1% Triton and blocked with 1% serum, 0.1% BSA in PBS for 30 minutes at room temperature. Samples were incubated with primary antibody (2ug/ml in blocking buffer) for 16 hours at 4°C. An Alexa Fluor® 488-conjugated Goat anti-rat IgG2a polyclonal was used as the secondary antibody (1/500). Total cells were stained using DAPI (blue).

Please note the data was generated from a different batch produced using hybridoma method.

This image is courtesy of an anonymous Abreview

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (phospho S28) antibody [HTA28] (AB10543)

Immunofluorescent analysis of 4% Paraformaldehyde fixed, 0.1% Triton X-100 permeabilized HeLa (human cervical adenocarcinoma epithelial cell) cells with ab10543 at 1/8000 dilution followed by ab150157 Goat Anti-Rat IgG H&L (Alexa Fluor® 488) antibody at 1/1000 (Green). ab179513 Anti-beta Tubulin rabbit monoclonal antibody was used as a counterstain at a 1/200 dilution followed by ab150088 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) preadsorbed at a 1/1000 dilution (Magenta).

Confocal image showing nuclear staining in HeLa cells (shown in green) at metaphase, the signal is decreased after λ Protein phosphatase treatment at 30℃ for 2 hours. The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (phospho S28) antibody [HTA28] (AB10543)

Immunofluorescent analysis of 4% Paraformaldehyde fixed, 0.1% Triton X-100 permeabilized HeLa (human cervical adenocarcinoma epithelial cell) cells with ab10543 at 1/8000 dilution followed by ab150157 Goat Anti-Rat IgG H&L (Alexa Fluor® 488) antibody at 1/1000 (Green). ab179513 Anti-beta Tubulin rabbit monoclonal antibody was used as a counterstain at a 1/200 dilution followed by ab150088 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) preadsorbed at a 1/1000 dilution (Magenta).

Confocal image showing nuclear staining in HeLa cells at metaphase (shown in green), and no staining at anaphase. The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (phospho S28) antibody [HTA28] (AB10543)

Immunocytochemical analysis of HeLa cells fixed and permeabilized with methanol followed by methanol : acetone. Histone H3 (phospho S28) was labeled with ab10543 at 5 μg/mL in ICC/IF (Green). The secondary antibody was a Goat Anti-Rat IgG-FITC conjugate. The nuclear counterstain was DAPI (Blue).

Please note the data was generated from a different batch produced using hybridoma method.

• ICC/IF

AbReview9421****

Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (phospho S28) antibody [HTA28] (AB10543)

ab10543 staining Histone H3 (phospho S28) in Mouse neural stem cells by ICC/IF (Immunocytochemistry/Immunofluorescence). Cells were fixed with paraformaldehyde and blocked with 4% serum for 1 hour at 20°C. Samples were incubated with primary antibody (1/500 in PBS + 4% serum) for 16 hours at 4°C. An Alexa Fluor® 546-conjugated anti-rat polyclonal was used as the secondary antibody (1/500). DAPI is stained blue

Please note the data was generated from a different batch produced using hybridoma method.

This image is courtesy of an anonymous Abreview

• WB

Lab

Western blot - Anti-Histone H3 (phospho S28) antibody [HTA28] (AB10543)

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

ab181602 was used as GAPDH loading control at a 1/2200000 dilution. ab201456 was used for total protein control.

All lanes:

Western blot - Anti-Histone H3 (phospho S28) antibody [HTA28] (ab10543) at 1/1000 dilution

Lane 1:

Untreated HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate (untreated membrane) at 20 µg

Lane 2:

HeLa treated with 100ng/ml nocodazole for 16 hours whole cell lysate (untreated membrane) at 20 µg

Lane 3:

Untreated HeLa whole cell lysate (Lambda Protein Phosphatase treated membrane) at 20 µg

Lane 4:

HeLa treated with 100ng/ml nocodazole for 16 hours whole cell lysate (Lambda Protein Phosphatase treated membrane) at 20 µg

Secondary

All lanes:

Western blot at 1/10000 dilution

Predicted band size: 15 kDa

false

Exposure time: 15s

• Dot

Lab

Dot Blot - Anti-Histone H3 (phospho S28) antibody [HTA28] (AB10543)

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

All lanes:

Dot Blot - Anti-Histone H3 (phospho S28) antibody [HTA28] (ab10543)

Lane 1:

Histone H3 non-phospho peptide

Lane 2:

Histone H3 (phospho S28) peptide

Secondary

All lanes:

Dot Blot - Goat Anti-Rat IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rat-igg-h-l-hrp-ab205720'>ab205720</a>) at 1/10000 dilution

Predicted band size: 15 kDa

false

Exposure time: 180s

• PepArr

Lab

Peptide Array - Anti-Histone H3 (phospho S28) antibody [HTA28] (AB10543)

ab10543 was tested in Peptide Array against 501 different modified and unmodified histone peptides; each peptide is printed on the array at six concentrations (each in triplicate). Circle area represents affinity between the antibody and a peptide : all antigen-containing peptides are displayed as red circles, all other peptides as blue circles. The affinity is calculated as the area under the curve when antibody binding values are plotted against the corresponding peptide concentration. Each circle area is normalized to the peptide with the strongest affinity. Goat Anti-Rat IgG, (H+L), Fluo 647nm conjugated was used as secondary at a 1/50000 dilution.

The complete dataset, including full list of all peptides and information on the position of each peptide in the diagram, is available under the Product Protocol section.