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9849
Phospho-Histone H3 (Mitotic Marker) Antibody Sampler Kit

Phospho-Histone H3 (Mitotic Marker) Antibody Sampler Kit #9849

Western Blotting Image 1

Western blot analysis of extracts from HeLa cells, either untreated or treated with nocodazole (100 ng/ml for 18 hours), using Phospho-Histone H3 (Ser10) (D2C8) XP® Rabbit mAb #3377 (upper) or Histone H3 Antibody #9715 (lower). Phospho-specificity of the antibody is shown by further treatment of the lysate with λ phosphatase.

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Western Blotting Image 2

Western blot analysis of lysates from HeLa, C6 and NIH/3T3 cells treated for 24 hours with or without nocodazole and also with or without λ phosphatase, using Phospho-Histone H3 (Thr11) Antibody #9764 (upper) or Histone H3 Antibody #9715 (lower).

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Western Blotting Image 3

Western blot analysis of lysates from CHO and HeLa cells either untreated or synchronized in metaphase by treatment with 100 ng/ml nocodazole for 4 h, followed by isolation of metaphase cells by mitotic shake-off. Blots were probed with Phospho-Histone H3 (Ser28) Antibody #9713 (upper) or Histone H3 Antibody #9715 (lower).

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Western Blotting Image 4

Western blot analysis of extracts from NIH/3T3 and HeLa cells, untreated or calyculin A and 20% FCS treated, using Phospho-Histone H3 (Thr3) Antibody (upper) or Histone H3 Antibody #9715 (lower).

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Western Blotting Image 5

Western blot analysis of extracts from various cell lines using Histone H3 (D1H2) XP® Rabbit mAb.

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Western Blotting Image 6

After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.

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Flow Cytometry Image 7

Flow cytometric analysis of Jurkat cells using Phospho-Histone H3 (Ser10) (D2C8) XP® Rabbit mAb versus propidium iodide (DNA content). The boxed population indicates Phospho-Histone H3 (Ser10) positive cells.

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Flow Cytometry Image 8

Flow cytometric analysis of untreated Jurkat cells, using Phospho-Histone H3 (Thr11) Antibody versus propidium iodide (DNA content). The boxed population indicates Phospho-Histone H3 (Thr11)-positive cells.

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Flow Cytometry Image 9

Flow cytometric analysis of untreated Jurkat cells, using Phospho-Histone H3 (Ser28) Antibody versus propidium iodide (DNA content). The box indicates phospho-Histone H3 positive cells.

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IHC-P (paraffin) Image 10

Immunohistochemical analysis of paraffin-embedded human colon carcinoma, showing nuclear localization, using Phospho-Histone H3 (Thr3) Antibody.

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IHC-P (paraffin) Image 11

Immunohistochemical analysis of paraffin-embedded human breast carcinoma using Histone H3 (D1H2) XP® Rabbit mAb.

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IF-IC Image 12

Confocal immunofluorescent analysis of HeLa cells using Phospho-Histone H3 (Ser10) (D2C8) XP® Rabbit mAb (green). Actin filaments have been labeled with DY-554 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

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IF-IC Image 13

Confocal immunofluorescent analysis showing positive signal in mitotic HeLa cells, using Phospho-Histone H3 (Thr11) Antibody (green). Actin filaments have been labeled with Alexa Fluor® 555 phalloidin (red).

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IF-IC Image 14

Confocal immunofluorescent analysis of C2C12 cells using Phospho-Histone H3 (Ser28) Antibody (green). Actin filaments have been labeled with Alexa Fluor® 555 phalloidin (red). Blue pseudocolor = DRAQ5™ (fluorescent DNA dye).

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IHC-P (paraffin) Image 15

Immunohistochemical analysis of paraffin-embedded MCF7 cells untreated (left) or nocodazole-treated (right), using Phospho-Histone H3 (Thr3) Antibody.

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Flow Cytometry Image 16

Flow cytometric analysis of human peripheral blood lymphocytes using Histone H3 (D1H2) XP® Rabbit mAb (blue) compared to Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (red). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 647 Conjugate) #4414 was used as a secondary antibody.

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IF-F Image 17

Confocal immunofluorescent analysis of postnatal day 1 rat brain using Phospho-Histone H3 (Ser28) Antibody (green). Blue pseudocolor = DRAQ5™ (fluorescent DNA dye).

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IHC-P (paraffin) Image 18

Immunohistochemical analysis of paraffin-embedded human tonsil untreated (left) or λ phosphatase-treated (right), using Phospho-Histone H3 (Thr3) Antibody.

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IF-IC Image 19

Confocal immunofluorescent analysis of HeLa cells using Histone H3 (D1H2) XP® Rabbit mAb (green) and β-Tubulin (9F3) Rabbit mAb (Alexa Fluor® 555 Conjugate) #2116 (red).

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Product Includes Quantity Applications Reactivity MW(kDa) Isotype
Phospho-Histone H3 (Ser10) (D2C8) XP® Rabbit mAb 3377 20 µl
  • WB
  • IF
  • F
H M R Mk Z 17 Rabbit IgG
Phospho-Histone H3 (Thr11) Antibody 9764 20 µl
  • WB
  • IP
  • IF
  • F
H M R 17 Rabbit 
Phospho-Histone H3 (Ser28) Antibody 9713 20 µl
  • WB
  • IP
  • IF
  • F
H M Hm Dm 17 Rabbit 
Phospho-Histone H3 (Thr3) Antibody 9714 20 µl
  • WB
  • IHC
H M R 17 Rabbit 
Histone H3 (D1H2) XP® Rabbit mAb 4499 20 µl
  • WB
  • IHC
  • IF
  • F
H M R Mk 17 Rabbit IgG
Anti-rabbit IgG, HRP-linked Antibody 7074 100 µl
  • WB
Goat 

The Phospho-Histone H3 (Mitotic Marker) Antibody Sampler Kit provides a fast and economical means of evaluating phosphorylation sites associated with mitosis on Histone H3. The kit contains enough primary and secondary antibodies to perform two Western blots.

All antibodies in the Phospho-Histone H3 Antibody Sampler Kit recognize Histone H3 only when modified at the indicated site.

Polyclonal antibodies are produced by immunizing animals with synthetic phosphopeptides corresponding to residues surrounding Thr3, Thr11 or Ser28 of human Histone H3. Antibodies are purified by protein A and peptide affinity chromatography. Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Ser10 of human histone H3.

Modulation of chromatin structure plays an important role in the regulation of transcription in eukaryotes. The nucleosome, made up of DNA wound around eight core histone proteins (two each of H2A, H2B, H3, and H4), is the primary building block of chromatin (1). The amino-terminal tails of core histones undergo various post-translational modifications, including acetylation, phosphorylation, methylation, and ubiquitination (2-5). These modifications occur in response to various stimuli and have a direct effect on the accessibility of chromatin to transcription factors and, therefore, gene expression (6). In most species, histone H2B is primarily acetylated at Lys5, 12, 15, and 20 (4,7). Histone H3 is primarily acetylated at Lys9, 14, 18, 23, 27, and 56. Acetylation of H3 at Lys9 appears to have a dominant role in histone deposition and chromatin assembly in some organisms (2,3). Phosphorylation at Ser10, Ser28, and Thr11 of histone H3 is tightly correlated with chromosome condensation during both mitosis and meiosis (8-10). Phosphorylation at Thr3 of histone H3 is highly conserved among many species and is catalyzed by the kinase haspin. Immunostaining with phospho-specific antibodies in mammalian cells reveals mitotic phosphorylation at Thr3 of H3 in prophase and its dephosphorylation during anaphase (11).

  1. Workman, J.L. and Kingston, R.E. (1998) Annu Rev Biochem 67, 545-79.
  2. Hansen, J.C. et al. (1998) Biochemistry 37, 17637-41.
  3. Strahl, B.D. and Allis, C.D. (2000) Nature 403, 41-5.
  4. Cheung, P. et al. (2000) Cell 103, 263-71.
  5. Bernstein, B.E. and Schreiber, S.L. (2002) Chem Biol 9, 1167-73.
  6. Jaskelioff, M. and Peterson, C.L. (2003) Nat Cell Biol 5, 395-9.
  7. Thorne, A.W. et al. (1990) Eur J Biochem 193, 701-13.
  8. Hendzel, M.J. et al. (1997) Chromosoma 106, 348-60.
  9. Goto, H. et al. (1999) J Biol Chem 274, 25543-9.
  10. Preuss, U. et al. (2003) Nucleic Acids Res 31, 878-85.
  11. Dai, J. et al. (2005) Genes Dev 19, 472-88.
For Research Use Only. Not For Use In Diagnostic Procedures.

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