Cell Signaling Technology
XP Monoclonal Antibody

Product Pathways - Lymphocyte Signaling

Phospho-HS1 (Tyr397) (D12C1) XP® Rabbit mAb #8714

Applications Reactivity Sensitivity MW (kDa) Isotype
W IF-IC F H (M) (R) Endogenous 80 Rabbit IgG

Applications Key:  W=Western Blotting  IF-IC=Immunofluorescence (Immunocytochemistry)  F=Flow Cytometry
Reactivity Key:  H=Human  M=Mouse  R=Rat
Species cross-reactivity is determined by western blot. Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Protocols

Specificity / Sensitivity

Phospho-HS1 (Tyr397) (D12C1) XP® Rabbit mAb recognizes endogenous levels of HS1 protein only when phosphorylated at Tyr397.

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Tyr405 of mouse HS1 protein. This site corresponds to Tyr397 of human HS1 protein.

Western Blotting

Western Blotting

Western blot analysis of extracts from Ramos cells, serum starved for 12 hours, then treated with anti-IgM antibody or λ phosphatase, using Phospho-HS1 (Tyr397) (D12C1) XP® Rabbit mAb (upper), or HS1 Antibody (Human Specific) #4503 (lower).

Flow Cytometry

Flow Cytometry

Flow cytometric analysis of Ramos cells, untreated (blue) or IgM-treated (green), using Phospho-HS1 (Tyr397) (D12C1) XP® Rabbit mAb.

IF-IC

IF-IC

Confocal immunofluorescent analysis of Ramos cells, untreated (left) or treated with IgM (12 μg/mL, 10 min, right) using Phospho-HS1 (Tyr397) (D12C1) XP® Rabbit mAb (green). Blue pseudocolor= DRAQ5® #4084 (fluorescent DNA dye).


Background

HS1 (HCLS1, LckBP1, p75) is a protein kinase substrate that is expressed only in tissues and cells of hematopoietic origin (1,2). HS1 contains four cortactin repeats and a single SH3 domain (2). This intracellular protein is phosphorylated following immune receptor activation, which promotes recruitment of HS1 to the immune synapse (3-5). Phosphorylation of HS1 is required to regulate actin dynamics and provide docking sites for many other signaling molecules, such as Vav1 and PLCγ1 (6). HS1 also plays an important role in platelet activation (7).

HS1 is rapidly phosphorylated at Tyr397 by Syk and/or Lyn kinases following immune receptor stimulation and thrombin-mediated platelet stimulation. This phosphorylation is an important step in cytoskeletal rearrangement and signaling complex formation (6-10).

  1. Kitamura, D. et al. (1989) Nucleic Acids Res 17, 9367-79.
  2. Kitamura, D. et al. (1995) Biochem Biophys Res Commun 208, 1137-46.
  3. Suzuki, H. et al. (1997) J Immunol 159, 5881-8.
  4. Hata, D. et al. (1994) Immunol Lett 40, 65-71.
  5. Yamanashi, Y. et al. (1993) Proc Natl Acad Sci USA 90, 3631-5.
  6. Gomez, T.S. et al. (2006) Immunity 24, 741-52.
  7. Kahner, B.N. et al. (2007) Blood 110, 2449-56.
  8. Yamanashi, Y. et al. (1997) J Exp Med 185, 1387-92.
  9. Hao, J.J. et al. (2004) J Biol Chem 279, 33413-20.
  10. Brunati, A.M. et al. (2005) J Biol Chem 280, 21029-35.

Application References

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Companion Products

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

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