Cell Signaling Technology

Product Pathways - Motif Antibodies

Phospho-(Ser) 14-3-3 Binding Motif Antibody #9601

Applications Reactivity Sensitivity Source
W IP IHC-P E-P All Endogenous Rabbit

Applications Key:  W=Western Blotting  IP=Immunoprecipitation  IHC-P=Immunohistochemistry (Paraffin)  E-P=ELISA (Peptide)
Reactivity Key: All=All species expected
Species cross-reactivity is determined by western blot.

Protocols

Specificity / Sensitivity

Phospho-(Ser) 14-3-3 Binding Motif Antibody binds peptides and proteins containing a motif composed of phospho-Ser with proline at the +2 position and arginine or lysine at the -3 position. Antibody binding is phospho-specific and largely independent of other surrounding amino acids. The antibody weakly cross-reacts with sequences containing phospho-Thr instead of phospho-Ser in this motif, and with sequences containing phospho-Ser surrounded by Phe at the +1 position and Arg/Lys at the -3 position. No cross-reactivity is observed with corresponding nonphosphorylated sequences or with other phospho-Thr/Ser/Tyr-containing motifs. By ELISA this antibody recognizes a wide range of peptides containing the 14-3-3 binding motif, and by 2D gel Western blot analysis it recognizes a large number of presumptive 14-3-3 binding proteins. (U.S. Patent No's.: 6,441,140; 6,982,318; 7,259,022; 7,344,714; U.S.S.N. 11,484,485; and all foreign equivalents.)

Source / Purification

Polyclonal antibodies are produced by immunizing animals with synthetic phospho-(Ser) 14-3-3-binding-motif containing peptides . Antibodies are purified by protein A and peptide affinity chromatography.

Western Blotting

Western Blotting

Western blot analysis of extracts from 293 cells transfected with Bad or Bad Ser112 and/or Ser136 mutant fusion proteins, using Phospho-(Ser) 14-3-3 Binding Motif Antibody (top), Phospho-Bad (Ser136) Antibody #9295 (middle) or Phospho-Bad (Ser112) Antibody #9291 (bottom).

Western Blotting

Western Blotting

Western blot analysis of extracts from Jurkat cells, untreated or calyculin A-treated (0.1 µM for 30 minutes), using Phospho-(Ser) 14-3-3 Binding Motif Antibody. Proteins were separated by 2-D electrophoresis prior to blotting.

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human breast carcinoma, showing staining of proteins containing phosphorylated 14-3-3 binding motifs, using Phospho-(Ser) 14-3-3 Binding Motif Antibody.


ELISA-Peptide

ELISA-Peptide

Phospho-(Ser) 14-3-3 Binding Motif Antibody ELISA Assay: Signal-to-noise ratio of phospho- versus nonphospho-14-3-3 binding motif peptides. (S* denotes phosphorylated serine.)

Background

The 14-3-3 proteins are a highly conserved family of proteins involved in the regulation of cell survival, apoptosis, proliferation and checkpoint control (1-5). Biological regulation by 14-3-3 is mediated through phosphorylation-dependent protein-protein interactions (6). Two different phospho-Ser-containing motifs are found within nearly all known 14-3-3 binding proteins (7). Motif 1 (Arg/Lys and Ser at positions -3 and -2, phospho-Ser at position 0, and Pro at position +2) is found in critical regulatory proteins including Bad, cdc25C, FKHRL1, PKC and c-Raf (5,7). Phospho-(Ser) 14-3-3 Binding Motif Polyclonal and (4E2) Monoclonal Antibodies provide powerful tools for the discovery and characterization of potential 14-3-3 binding proteins containing this motif and for high throughput drug discovery.

  1. Aitken, A. (1995) Trends Biochem Sci 20, 95-7.
  2. Zha, J. et al. (1996) Cell 87, 619-28.
  3. Piwnica-Worms, H. (1999) Nature 401, 535, 537.
  4. Tzivion, G. et al. (1998) Nature 394, 88-92.
  5. Xing, H. et al. (2000) EMBO J 19, 349-58.
  6. Muslin, A.J. et al. (1996) Cell 84, 889-97.
  7. Yaffe, M.B. et al. (1997) Cell 91, 961-71.

Application References

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Companion Products

License/Use Restrictions: Use of CST Motif Antibodies within certain methods (e.g., U.S. Patent No.'s 7,198,896 & 7,300,753) may require a license from CST. For information regarding academic licensing terms please have your technology transfer office contact CST Legal Department at CST_ip@cellsignal.com. For information regarding commercial licensing terms please contact CST Pharma Services Department at ptmscan@cellsignal.com.

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

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