Nocodazole treated HeLa cells were subjected to SDS PAGE followed by western blot with 82828-10-RR (Phospho-Histone H3 (Ser10) antibody) at dilution of 1:2000 incubated at room temperature for 1.5 hours. The membrane was stripped and reblotted with Alpha Tubulin Monoclonal antibody (66031-1-Ig) as loading control.
Nocodazole treated HeLa cells were subjected to SDS PAGE followed by western blot with 82828-10-RR (Phospho-Histone H3 (Ser10) antibody) at dilution of 1:2000 incubated at room temperature for 1.5 hours. The membrane was stripped and reblotted with Alpha Tubulin Monoclonal antibody (66031-1-Ig) as loading control.
WB analysis of NIH/3T3 using 82828-10-RR
Nocodazole treated and untreated NIH/3T3 cells were subjected to SDS PAGE followed by western blot with 82828-10-RR (Phospho-Histone H3 (Ser10) antibody) at dilution of 1:2000 incubated at room temperature for 1.5 hours. The membrane was stripped and reblotted with GAPDH Monoclonal antibody (60004-1-Ig) as loading control.
Nocodazole treated and untreated NIH/3T3 cells were subjected to SDS PAGE followed by western blot with 82828-10-RR (Phospho-Histone H3 (Ser10) antibody) at dilution of 1:2000 incubated at room temperature for 1.5 hours. The membrane was stripped and reblotted with GAPDH Monoclonal antibody (60004-1-Ig) as loading control.
WB analysis of HSC-T6 using 82828-10-RR
Calyculin A treated and untreated HSC-T6 cells were subjected to SDS PAGE followed by western blot with 82828-10-RR (Phospho-Histone H3 (Ser10) antibody) at dilution of 1:2000 incubated at room temperature for 1.5 hours. The membrane was stripped and reblotted with GAPDH Monoclonal antibody (60004-1-Ig) as loading control.
Calyculin A treated and untreated HSC-T6 cells were subjected to SDS PAGE followed by western blot with 82828-10-RR (Phospho-Histone H3 (Ser10) antibody) at dilution of 1:2000 incubated at room temperature for 1.5 hours. The membrane was stripped and reblotted with GAPDH Monoclonal antibody (60004-1-Ig) as loading control.
IHC staining of mouse brain using 82828-10-RR
Immunohistochemical analysis of paraffin-embedded mouse brain tissue slide using 82828-10-RR (Phospho-Histone H3 (Ser10) antibody) at dilution of 1:1000 (under 10x lens). Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0).
Immunofluorescent analysis of (4% PFA) fixed HepG2 cells using Phospho-Histone H3 (Ser10) antibody (82828-10-RR, Clone: 242921B1 ) at dilution of 1:800 and CoraLite®488-Conjugated Goat Anti-Rabbit IgG(H+L) (SA00013-2), CL594-Phalloidin (red).
ChIP experiment of HeLa using 82828-10-RR
Chromatin was prepared from HeLa cells either non-treated or treated with Nocodazole (100 ng/ml, 16 h). Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 20 µg of cross-linked chromatin, 5 µg of Phospho-Histone H3 (Ser10) (82828-10-RR) or 5 ug of Normal Rabbit IgG (98136-1-RR), and 20 µl of Protein A Magarose Beads. The immunoprecipitated DNA was quantified by real-time PCR.
Chromatin was prepared from HeLa cells either non-treated or treated with Nocodazole (100 ng/ml, 16 h). Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 20 µg of cross-linked chromatin, 5 µg of Phospho-Histone H3 (Ser10) (82828-10-RR) or 5 ug of Normal Rabbit IgG (98136-1-RR), and 20 µl of Protein A Magarose Beads. The immunoprecipitated DNA was quantified by real-time PCR.
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nocodazole treated HeLa cells, Calyculin A treated HSC-T6 cells, nocodazole NIH/3T3 cells
Positive IHC detected in
mouse brain tissue Note: suggested antigen retrieval with TE buffer pH 9.0; (*) Alternatively, antigen retrieval may be performed with citrate buffer pH 6.0
Positive IF/ICC detected in
HepG2 cells
Positive ChIP-qPCR detected in
Nocodazole (100 ng/ml, 16 h) HeLa cells
Recommended dilution
Application
Dilution
Western Blot (WB)
WB : 1:1000-1:4000
Immunohistochemistry (IHC)
IHC : 1:500-1:2000
Immunofluorescence (IF)/ICC
IF/ICC : 1:400-1:1600
CHIP-QPCR
CHIP-QPCR : 1:10-1:100
It is recommended that this reagent should be titrated in each testing system to obtain optimal results.
Sample-dependent, Check data in validation data gallery.
Product Information
82828-10-RR targets Phospho-Histone H3 (Ser10) in WB, IHC, IF/ICC, ELISA, ChIP-qPCR applications and shows reactivity with human, mouse, rat samples.
PBS with 0.02% sodium azide and 50% glycerol, pH 7.3.
Storage Conditions
Store at -20°C. Stable for one year after shipment. Aliquoting is unnecessary for -20oC storage. 20ul sizes contain 0.1% BSA.
Background Information
Phospho-histone-H3 (PHH3) is a core histone protein, which in its phosphorylated state forms the principal constituents of eukaryotic chromatin, with histone H3 being phosphorylated at serine (Ser) 10 or Ser28 as well as its phosphorylation of Ser10 being strongly correlated with the late G2 to M-phase transition in mammalian mitotic cells. On the basis of previous research, a few cell line- and animal model-based researches have displayed an increase in phosphorylation of histone H3 at Ser10 (H3S10ph), the only histone marker that is involved in carcinogenesis and cellular transformation. Histone H3 phosphorylation on serine-10 is specific to mitosis and phosphorylated histone H3 (PHH3) proliferation markers (as counts defined per area or as indices defined per cell numbers) are increasingly being used to evaluate proliferation in various tumors.
Protocols
Product Specific Protocols
WB protocol for Phospho-Histone H3 (Ser10) antibody 82828-10-RR
Nocodazole treated HeLa cells were subjected to SDS PAGE followed by western blot with 82828-10-RR (Phospho-Histone H3 (Ser10) antibody) at dilution of 1:2000 incubated at room temperature for 1.5 hours. The membrane was stripped and reblotted with Alpha Tubulin Monoclonal antibody (66031-1-Ig) as loading control.
WB analysis of NIH/3T3 using 82828-10-RR
Nocodazole treated and untreated NIH/3T3 cells were subjected to SDS PAGE followed by western blot with 82828-10-RR (Phospho-Histone H3 (Ser10) antibody) at dilution of 1:2000 incubated at room temperature for 1.5 hours. The membrane was stripped and reblotted with GAPDH Monoclonal antibody (60004-1-Ig) as loading control.
WB analysis of HSC-T6 using 82828-10-RR
Calyculin A treated and untreated HSC-T6 cells were subjected to SDS PAGE followed by western blot with 82828-10-RR (Phospho-Histone H3 (Ser10) antibody) at dilution of 1:2000 incubated at room temperature for 1.5 hours. The membrane was stripped and reblotted with GAPDH Monoclonal antibody (60004-1-Ig) as loading control.
IHC Figures
IHC staining of mouse brain using 82828-10-RR
Immunohistochemical analysis of paraffin-embedded mouse brain tissue slide using 82828-10-RR (Phospho-Histone H3 (Ser10) antibody) at dilution of 1:1000 (under 10x lens). Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0).
IF/ICC Figures
IF Staining of HepG2 using 82828-10-RR
Immunofluorescent analysis of (4% PFA) fixed HepG2 cells using Phospho-Histone H3 (Ser10) antibody (82828-10-RR, Clone: 242921B1 ) at dilution of 1:800 and CoraLite®488-Conjugated Goat Anti-Rabbit IgG(H+L) (SA00013-2), Beta Tubulin antibody (66240-1-Ig, Clone: 1D4A4, red).
IF Staining of HepG2 using 82828-10-RR
Immunofluorescent analysis of (4% PFA) fixed HepG2 cells using Phospho-Histone H3 (Ser10) antibody (82828-10-RR, Clone: 242921B1 ) at dilution of 1:800 and CoraLite®488-Conjugated Goat Anti-Rabbit IgG(H+L) (SA00013-2), Beta Tubulin antibody (66240-1-Ig, Clone: 1D4A4, red).
IF Staining of HepG2 using 82828-10-RR
Immunofluorescent analysis of (4% PFA) fixed HepG2 cells using Phospho-Histone H3 (Ser10) antibody (82828-10-RR, Clone: 242921B1 ) at dilution of 1:800 and CoraLite®488-Conjugated Goat Anti-Rabbit IgG(H+L) (SA00013-2), CL594-Phalloidin (red).
CHIP-QPCR Figures
ChIP experiment of HeLa using 82828-10-RR
Chromatin was prepared from HeLa cells either non-treated or treated with Nocodazole (100 ng/ml, 16 h). Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 20 µg of cross-linked chromatin, 5 µg of Phospho-Histone H3 (Ser10) (82828-10-RR) or 5 ug of Normal Rabbit IgG (98136-1-RR), and 20 µl of Protein A Magarose Beads. The immunoprecipitated DNA was quantified by real-time PCR.
The species listed in Tested Reactivity are in-house verified and applicable species. For unlisted species, please refer to the homology analysis of the immunogen sequence and related species. For rabbit polyclonal antibodies, homology >70% is recommended. For mouse monoclonal antibodies and rabbit recombinant antibodies, homology >90% is recommended. Generally, the higher the homology, the greater the applicability. However, there will be certain differences in protein expression in different species, tissues or cells. Therefore, the homology analysis results are for reference only and do not serve as a guarantee.
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