Cell Signaling Technology
XP Monoclonal Antibody

Product Pathways - Chromatin Regulation / Epigenetics

Bmi1 (D20B7) XP® Rabbit mAb #6964

Applications Reactivity Sensitivity MW (kDa) Isotype
W IP IHC-P IF-IC ChIP H Mk Endogenous 41, 43 Rabbit IgG

Applications Key:  W=Western Blotting  IP=Immunoprecipitation  IHC-P=Immunohistochemistry (Paraffin)  IF-IC=Immunofluorescence (Immunocytochemistry)  ChIP=Chromatin IP
Reactivity Key:  H=Human  Mk=Monkey
Species cross-reactivity is determined by western blot. Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Protocols

Specificity / Sensitivity

Bmi1 (D20B7) XP® Rabbit mAb recognizes endogenous levels of total Bmi1 protein.

Source / Purification

Monoclonal antibody is produced by immunizing animals with recombinant protein specific to the carboxy terminus of human Bmi1 protein.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using Bmi1 (D20B7) XP® Rabbit mAb.

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human lung carcinoma using Bmi1 (D20B7) XP® Rabbit mAb.

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human lymphoma using Bmi1 (D20B7) XP® Rabbit mAb.


IF-IC

IF-IC

Confocal immunofluorescent analysis of COS-7 cells using Bmi1 (D20B7) XP® Rabbit mAb (green). Actin filaments were labeled with DY-554 phalloidin (red).

Chromatin IP

Chromatin IP

Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 NCCIT cells and either 10 µl of Bmi1 (D20B7) XP® Rabbit mAb or 2 µl of Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human HoxA1 Intron 1 Primers #7707, SimpleChIP® Human HoxA2 Promoter Primers #5517, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin (equivalent to one).

Background

The polycomb group (PcG) of proteins contributes to the maintenance of cell identity, stem cell self-renewal, cell cycle regulation, and oncogenesis by maintaining the silenced state of genes that promote cell lineage specification, cell death, and cell-cycle arrest (1-4). PcG proteins exist in two complexes that cooperate to maintain long-term gene silencing through epigenetic chromatin modifications. The first complex, EED-EZH2, is recruited to genes by DNA-binding transcription factors and methylates histone H3 on Lys27. This histone methyl-transferase activity requires the Ezh2, Eed, and Suz12 subunits of the complex (5). Histone H3 methylation at Lys27 facilitates the recruitment of the second complex, PRC1, which ubiquitinylates histone H2A on Lys119 (6). Bmi1 is a component of the PRC1 complex, which together with Ring1 strongly enhances the E3 ubiquitin ligase activity of the Ring2 catalytic subunit (7). Bmi1 plays an important role in the regulation of cell proliferation and senescence through repression of the p16 INK4A and p19 ARF genes and is required for maintenance of adult hematopoietic and neural stem cells (3,4,8-10).

  1. Boyer, L.A. et al. (2006) Nature 441, 349-53.
  2. Lee, T.I. et al. (2006) Cell 125, 301-13.
  3. Park, I.K. et al. (2003) Nature 423, 302-5.
  4. Molofsky, A.V. et al. (2003) Nature 425, 962-7.
  5. Cao, R. and Zhang, Y. (2004) Mol Cell 15, 57-67.
  6. Wang, H. et al. (2004) Nature 431, 873-8.
  7. Cao, R. et al. (2005) Mol Cell 20, 845-54.
  8. Molofsky, A.V. et al. (2005) Genes Dev 19, 1432-7.
  9. Jacobs, J.J. et al. (1999) Nature 397, 164-8.
  10. Jacobs, J.J. et al. (1999) Genes Dev 13, 2678-90.

Application References

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Companion Products

For Research Use Only. Not For Use In Diagnostic Procedures.

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