Cell Signaling Technology

Product Pathways - Motif Antibodies

Phospho-(Ser/Thr) Akt Substrate Antibody #9611

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

Applications Key:  W=Western Blotting  IP=Immunoprecipitation  IHC-P=Immunohistochemistry (Paraffin)  E-P=ELISA (Peptide)
Reactivity Key: All=All species expected

Specificity / Sensitivity

Phospho-(Ser/Thr) Akt Substrate Antibody preferentially recognizes peptides and proteins containing phospho-Ser/Thr preceded by Lys/Arg at positions -5 and -3, in a manner largely independent of other surrounding amino acids. Some cross-reactivity is observed for peptides that contain phospho-Ser/Thr preceded by Arg/Lys at positions -3 and -2. No cross-reactivity is observed with the corresponding nonphosphorylated sequences or with other phospho-Ser/Thr-containing motifs. By ELISA, the antibody recognizes a wide range of phosphorylated Akt substrate peptides, and, by 2-D gel Western blot analysis, it recognizes a large number of proteins presumed to be Akt substrates. (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 rabbits with phospho-Akt substrate peptides (KLH-coupled). Antibodies are purified by protein A and peptide affinity chromatography.

Western Blotting

Western Blotting

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

Western Blotting

Western Blotting

Western blot analysis of extracts from A431 cells, untreated or calyculin A-treated, using Phospho-(Ser/Thr) Akt Substrate Antibody.

Western Blotting

Western Blotting

Western blot analysis of extracts from C2C12 cells, untreated or insulin-treated (100 nM for 15 minutes prior to lysis), using Phospho-(Ser/Thr) Akt Substrate Antibody.


IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human breast carcinoma control (left) or lambda phosphatase-treated (right), using Phospho-(Ser/Thr) Akt Substrate Antibody.

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human colon carcinoma, using Phospho-(Ser/Thr) Akt Substrate Antibody.

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human lung carcinoma, using Phospho-(Ser/Thr) Akt Substrate Antibody.


ELISA-Peptide

ELISA-Peptide

Phospho-(Ser/Thr) Akt Substrate Antibody DELFIA® Assay: Signal-to-noise ratio of phospho- versus nonphospho-peptides. (T* and S* denote phosphorylated threonine and serine.) (DELFIA® is a registered trademark of PerkinElmer, Inc.)

Background

An important class of kinases, referred to as Arg-directed kinases or AGC-family kinases, includes cAMP-dependent protein kinase (PKA), cGMP-dependent protein kinase (PKG), protein kinase C, Akt and RSK. These kinases share a substrate specificity characterized by Arg at position -3 relative to the phosphorylated Ser or Thr (1,2).Akt plays a central role in mediating critical cellular responses including cell growth and survival, angiogenesis and transcriptional regulation (3-5). While a number of Akt substrates including GSK-3, Bad and caspase-9 are known, many important substrates await discovery. Akt phosphorylates substrates only at serine/threonine in a conserved motif characterized by arginine at positions -5 and -3 (6).Phospho-Akt substrate-specific antibodies from Cell Signaling Technology are powerful tools for investigating the regulation of phosphorylation by Akt and other Arg-directed kinases, as well as for high throughput kinase drug discovery.

  1. Montminy, M. et al. (1997) Annu. Rev. Biochem. 66, 807-822.
  2. Pearson, R.B. and Kemp, B.E. (1991) Methods Enzymol. 200, 62-82.
  3. Marte, B.M. and Downward, J. (1997) Trends Biochem. Sci. 22, 355-358.
  4. Jiang, B. H. et al. (2000) Proc. Natl. Acad. Sci. USA 97, 1749-1753.
  5. Scheid, M.P. and Woodgett, J.R. (2000) Curr. Biol. 10, R191-R194.
  6. Alessi, D. R. et al. (1996) FEBS Lett. 399, 333-338.

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 commerical licensing terms please contact CST Business Development at cbunker@cellsignal.com.

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