Revision 1
Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:

WB, IP, E-P

REACTIVITY:

All

SENSITIVITY:

Endogenous

MW (kDa):

Source/Isotype:

Mouse IgG2a

Product Information

Product Usage Information

Application Dilution
Western Blotting 1:1000
Immunoprecipitation 1:50
Peptide ELISA (DELFIA) 1:1000

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

Specificity / Sensitivity

Phospho-(Ser/Thr) PDK1 Docking Motif (18A2) Mouse mAb detects phosphorylated serine or threonine that is surrounded by tyrosine or phenylalanine at the -1 and +1 positions and phenylalanine at the -4 position. It also recognizes peptides containing lysine instead of phenylalanine at the -4 position. This antibody does not cross-react with the nonphosphorylated PDK1 docking motifs or with other phosphorylated motifs. This antibody detects endogenous levels of phosphorylated proteins containing PDK1 docking motif, including phospho-Akt.

Species Reactivity:

All Species Expected

Source / Purification

Monoclonal antibody is produced by immunizing animals with peptides containing the PDK1 docking motif.

Background

A hallmark of signal transduction pathways is the reversible phosphorylation of serine and threonine residues within specific sequences, or motifs, in target proteins. Specific signaling motifs include not only sequences that are recognized by protein kinases (1), but also those that are recognized by phosphorylation-dependent binding proteins such as 14-3-3 (2). These modular phosphoprotein interacting domains are critical elements in modulating, directing and amplifying intracellular communications. CST has pioneered the development of phospho-motif specific antibodies, which are invaluable tools for probing the complexity of phospho-regulatory pathways.
Many critical protein kinases can be regulated by phosphorylation at a specific serine or threonine in a hydrophobic motif (3). For example, Akt, a kinase that regulates cell survival, is activated by phosphorylation at Ser473, a site preceded by Phe at -4 and -1 and followed by Tyr at +1 (4). RSK2, p70 S6 kinase and certain PKC isoforms also contain a similar consensus phosphorylation motif. Phosphorylation of these motifs is required for binding to 3-phosphoinositide-dependent kinase 1 (PDK1) (5-7). Phospho-(Ser/Thr) PDK1 Docking Motif (18A2) Monoclonal Antibody is a powerful tool for the characterization of phosphorylated PDK1 docking motifs and the identification of new proteins with PDK1 docking motifs.

  1. Pinna, L.A. and Ruzzene, M. (1996) Biochim Biophys Acta 1314, 191-225.
  2. Yaffe, M.B. and Elia, A.E. (2001) Curr Opin Cell Biol 13, 131-8.
  3. Vanhaesebroeck, B. and Alessi, D.R. (2000) Biochem J 346 Pt 3, 561-76.
  4. Alessi, D.R. et al. (1996) EMBO J 15, 6541-51.
  5. Frödin, M. et al. (2000) EMBO J 19, 2924-34.
  6. Balendran, A. et al. (1999) J Biol Chem 274, 37400-6.
  7. Balendran, A. et al. (2000) J Biol Chem 275, 20806-13.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IP: Immunoprecipitation E-P: Peptide ELISA (DELFIA)

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

Limited Uses

Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

Revision 1
#9634

Phospho-(Ser/Thr) PDK1 Docking Motif (18A2) Mouse mAb

Western Blotting Image 1: Phospho-(Ser/Thr) PDK1 Docking Motif (18A2) Mouse mAb Expand Image
Western blot analysis of extracts from A431 cells, untreated or calyculin A-treated (0.1 µM for 30 minutes prior to lysis), using Phospho-(Ser/Thr) PDK1 Docking Motif (18A2) Mouse mAb.
Immunoprecipitation Image 1: Phospho-(Ser/Thr) PDK1 Docking Motif (18A2) Mouse mAb Expand Image
Immunoprecipitation of extracts from Jurkat cells, untreated or treated with 0.1nM Calyculin A for 30 minutes prior to lysis, using Phospho-(Ser/Thr) Docking Motif (18A2) Mouse mAb. Lanes 1 and 2 are 10% input, lanes 3 and 4 are Protein A Agarose Beads #9863 control, and lanes 5 and 6 are Phospho-(Ser/Thr) Docking Motif (18A2) Mouse mAb. Western blot analysis was performed using Phospho-(Ser/Thr) Docking Motif (18A2) Mouse mAb.
Product Image 1: Phospho-(Ser/Thr) PDK1 Docking Motif (18A2) Mouse mAb Expand Image
Phospho-(Ser/Thr) PDK1 Docking Motif (18A2) Mouse mAb ELISA Assay: Signal-to-noise ratio of phospho- versus nonphospho-peptides. (T* and S* denote phosphorylated threonine and serine.)