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9768
B Cell Signaling Antibody Sampler Kit

B Cell Signaling Antibody Sampler Kit #9768

Western Blotting Image 1

Western blot analysis of extracts NIH/3T3 cells, serum-starved or treated with human Platelet-Derived Growth Factor BB hPDGF-BB #8912 (100 ng/ml, 15 min), using Phospho-Src Family (Tyr416) (D49G4) Rabbit mAb (upper) or Src (36D10) Rabbit mAb #2109 (lower).

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Western Blotting Image 2

Western blot analysis of extracts from Ramos cells, untreated or treated with anti-IgM, using Phospho-Syk (Tyr525/526) (C87C1) Rabbit mAb (upper) or Syk Antibody #2712 (lower).

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Western Blotting Image 3

Western blot analysis of extracts from Ramos cells, untreated or treated with anti-human IgM (12 µg/ml for 10 minutes), using Phospho-Btk (Tyr223) Antibody (upper) or Btk (C82B8) Rabbit mAb #3533 (lower).

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Western Blotting Image 4

Western blot analysis of extracts from untreated or anti-human IgM treated Ramos cells or lamda phosphatase-treated Ramos cell lysates, using Phospho-PLCgamma2 (Tyr759) Antibody (upper) or PLCgamma2 Antibody #3872 (lower).

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Western Blotting Image 5

Western blot analysis of SDS extracts from control or anti-human IgM treated (12 µg/ml for 2 minutes) Ramos cells using Phospho-BLNK (Tyr96) Antibody.

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Western Blotting Image 6

Western blot analysis of SDS extracts from untreated or anti-human IgM treated (12 µg/ml for 2 minutes) Ramos cells using Phospho-CD19 (Tyr531) Antibody (upper) and control CD19 Antibody #3574 (lower).

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Western Blotting Image 7

Western blot analysis of extracts from Ramos cells untreated or treated with anti-human IgM (12 µg/ml for 10 minutes), using Phospho-CD79A (Tyr182) Antibody (upper) or CD79A Antibody #3351 (lower).

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Western Blotting Image 8

Western blot analysis of extracts from Raji and Daudi cell lines using CD79A Antibody.

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Western Blotting Image 9

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 10

Flow cytometric analysis of Ramos cells, untreated (blue) or treated with anti-IgM (green), using Phospho-Syk(Tyr525/526) (C87C1) Rabbit mAb. Anti-rabbit IgG (H+L), F(ab')2 Fragment (PE Conjugate) #8885 was used as a secondary antibody.

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Flow Cytometry Image 11

Flow cytometric analysis of Ramos cells, untreated (blue) or IgM-treated (green), using Phospho-CD79A (Tyr182) Antibody.

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IF-IC Image 12

Confocal immunofluorescent analysis of Ramos (left) and Jurkat cells (right) using CD79A Antibody (green). Blue pseudocolor= DRAQ5® #4084 (fluorescent DNA dye).

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IF-IC Image 13

Confocal immunofluorescent analysis of Ramos cells, serum-starved (overnight; left) or IgM-treated (12 ug/ml, 2 minutes; right), using Phospho-Syk (Tyr525/526) (C87C1) Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

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IF-IC Image 14

Confocal immunofluorescent analysis of Ramos cells, untreated (left) or IgM-treated (middle), and Jurkat cells (right) using Phospho-CD79A (Tyr182) Antibody (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

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Product Includes Quantity Applications Reactivity MW(kDa) Isotype
Phospho-Src Family (Tyr416) (D49G4) Rabbit mAb 6943 20 µl
  • WB
  • IP
H M R Mk 60 Rabbit IgG
Phospho-Syk (Tyr525/526) (C87C1) Rabbit mAb 2710 20 µl
  • WB
  • IP
  • IF
  • F
H 72 Rabbit IgG
Phospho-Btk (Tyr223) Antibody 5082 20 µl
  • WB
H 77 Rabbit 
Phospho-PLCγ2 (Tyr759) Antibody 3874 20 µl
  • WB
H M 150 Rabbit 
Phospho-BLNK (Tyr96) Antibody 3601 20 µl
  • WB
  • IP
H 68, 70 Rabbit 
Phospho-CD19 (Tyr531) Antibody 3571 20 µl
  • WB
  • IP
H 95 Rabbit 
Phospho-CD79A (Tyr182) Antibody 5173 20 µl
  • WB
  • IP
  • IF
  • F
H 45-55 Rabbit 
CD79A Antibody 3351 20 µl
  • WB
  • IF
H 45-55 Rabbit 
Anti-rabbit IgG, HRP-linked Antibody 7074 100 µl
  • WB
Goat 

The B Cell Signaling Antibody Sampler Kit provides an economical means to examine key signaling proteins commonly associated with B cell activation. The provided antibodies allow monitoring of both total protein levels and the phosphorylation state. The kit includes enough primary and secondary antibody to perform two western mini-blot experiments.

All antibodies contained in this kit detect endogenous levels of their respective target protein.

Polyclonal antibodies are produced by immunizing animals with a synthetic peptide and are purified by protein A and peptide affinity chromatography. Monoclonal antibodies are produced by immunizing animals with recombinant human proteins or synthetic peptides.

Antigen receptors found on the surface of B cells contain a heterodimeric signaling component composed of CD79A and CD79B, also known as Ig α and Ig ß, respectively. Presence of this receptor complex is essential for B-cell development and function. Antigen binding precedes formation of the CD79A and CD79B heterodimer and subsequent activation of receptor associated kinases. Tyr182 of mouse CD79A (corresponding to Tyr188 of human CD79A) is one of two key tyrosine residues in the immunoreceptor tyrosine-based activation motif (ITAM) of CD79A that are phosphorylated by Src family kinases (e.g., Lyn, Blk), and play a critical role in modulating signal transduction following immune receptor activation.

Syk is a protein tyrosine kinase that plays an important role in intracellular signal transduction in hematopoietic cells (1-3). Syk interacts with immunoreceptor tyrosine-based activation motifs (ITAMs) located in the cytoplasmic domains of immune receptors (4). It couples the activated immunoreceptors to downstream signaling events that mediate diverse cellular responses, including proliferation, differentiation, and phagocytosis (4). There is also evidence that Syk plays a role in nonimmune cells; Syk is a potential tumor suppressor in human breast carcinomas (5). Tyrosine 525 and 526 are located in the activation loop of the Syk kinase domain, and phosphorylation of Tyr525/526 of human Syk (equivalent to the Tyr519/520 of mouse Syk) is essential for Syk function (6).

Lyn, one of the Src family members, is predominantly expressed in hematopoietic cells (7). Two tyrosine residues have been reported to play a crucial role in the regulation of protein tyrosine kinases of the Src family. Autophosphorylation of Tyr396 (equivalent to Tyr416 of Src), located in the catalytic domain, correlates with enzyme activation. Csk-mediated phosphorylation of the carboxy-terminal Tyr507 (equivalent to Tyr527 of Src) inactivates the kinase. Tyrosine phosphorylation and activation of Lyn occurs upon association with cell surface receptors such as the B cell Ag receptor (BCR) and CD40 (8-10).

Bruton's tyrosine kinase (Btk) is a member of the Btk/Tec family of cytoplasmic tyrosine kinases. Btk plays an important role in B cell development (11,12). Activation of B cells by various ligands is accompanied by Btk membrane translocation mediated by its PH domain binding to phosphatidylinositol-3,4,5-trisphosphate (13-15). The membrane-located Btk is active and associated with transient phosphorylation of two tyrosine residues, Tyr551 and Tyr223. Tyr551 in the activation loop is transphosphorylated by the Src family tyrosine kinase, leading to autophosphorylation at Tyr223 within the SH3 domain, which is necessary for full activation (16,17).

CD19 is a 95 kDa coreceptor that amplifies the signaling cascade in B cells (18). On the B cell surface, CD19 associates with CD21, CD81, and Leu-13 to exert its function. The cytoplasmic tail of CD19 has nine conserved tyrosine residues playing critical roles in CD19-mediated function by coupling signaling molecules to the receptor (18). After BCR or CD19 ligation, Tyr531 and Tyr500 of CD19 are progressively phosphorylated. This phosphorylation enables the coupling of PI3 kinase and Src family tyrosine kinase to CD19 and activates the PI3K and Src signaling pathways (19,20).

B cell linker protein (BLNK), also known as SLP-65 or BASH, is an adaptor molecule that plays key roles in B cell activation and B cell antigen receptor (BCR) engagement. BLNK acts at the interface between BCR-associated Syk and downstream signaling cascades

Phosphoinositide-specific phospholipase C (PLC) plays a significant role in transmembrane signaling. PLCgamma2 is engaged in antigen-dependent signaling in B cells. Phosphorylation by Btk or Lck at tyrosines 753, 759, 1197 and 1217 is correlated with PLCgamma2 activity.

  1. Cheng, A.M. and Chan, A.C. (1997) Curr Opin Immunol 9, 528-33.
  2. Chu, D.H. et al. (1998) Immunol Rev 165, 167-80.
  3. Yamanashi, Y. et al. (1989) Proc. Natl. Acad. Sci. USA 86, 6538-6542.
  4. Khan, W.N. (2001) Immunol Res 23, 147-56.
  5. Tedder, T.F. et al. (1997) Immunity 6, 107-18.
  6. Kurosaki, T. (1997) Curr Opin Immunol 9, 309-18.
  7. Yamanashi, Y. et al. (1991) Science 251, 192-194.
  8. Lewis, C.M. et al. (2001) Curr Opin Immunol 13, 317-25.
  9. Buhl, A.M. and Cambier, J.C. (1999) J Immunol 162, 4438-46.
  10. Burkhardt, A.L. et al. (1991) Proc. Natl. Acad. Sci. USA 88, 7410-7414.
  11. Salim, K. et al. (1996) EMBO J 15, 6241-50.
  12. Fujimoto, M. et al. (2000) Immunity 13, 47-57.
  13. Turner, M. et al. (2000) Immunol Today 21, 148-54.
  14. Ren, C.L. et al. (1994) J. Exp. Med. 179, 673-680.
  15. Rameh, L.E. et al. (1997) J Biol Chem 272, 22059-66.
  16. Coopman, P.J. et al. (2000) Nature 406, 742-7.
  17. Várnai, P. et al. (1999) J Biol Chem 274, 10983-9.
  18. Rawlings, D.J. et al. (1996) Science 271, 822-5.
  19. Park, H. et al. (1996) Immunity 4, 515-25.
  20. Zhang, J. et al. (2000) J Biol Chem 275, 35442-7.
Entrez-Gene Id
29760 , 973 , 2534 , 3055 , 3932 , 4067 , 5336 , 6714 , 7525
Swiss-Prot Acc.
Q8WV28 , P11912 , P06241 , P08631 , P06239 , P07948 , P16885 , P12931 , P07947
For Research Use Only. Not For Use In Diagnostic Procedures.

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
U.S. Patent No. 7,429,487, foreign equivalents, and child patents deriving therefrom.

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