Cell Signaling Technology

Product Pathways - Translational Control

p70 S6 Kinase Substrates Antibody Sampler Kit #2903

Kit Includes Quantity Applications Reactivity MW (kDa) Isotype
Phospho-S6 Ribosomal Protein (Ser235/236) (D57.2.2E) XP® Rabbit mAb #4858 40 µl W IHC-P IHC-F IF-IC F H M R Mk Mi Sc (C) (Pg) 32 Rabbit IgG
Phospho-S6 Ribosomal Protein (Ser240/244) (D68F8) XP® Rabbit mAb #5364 40 µl W IP IHC-P IF-IC F H M R Mk (C) (Pg) 32 Rabbit IgG
Phospho-eIF4B (Ser422) Antibody #3591 40 µl W H M R Mk 80 Rabbit
Phospho-eEF2k (Ser366) Antibody #3691 40 µl W IP H R Mk 105 Rabbit
Phospho-p70 S6 Kinase (Thr389) (108D2) Rabbit mAb #9234 40 µl W H M R Mk (C) 70, 85 Rabbit IgG
p70 S6 Kinase (49D7) Rabbit mAb #2708 40 µl W IHC-P H M R Mk 70, 85 Rabbit IgG
Anti-rabbit IgG, HRP-linked Antibody #7074 100 µl Goat

Applications Key:  W=Western Blotting  IP=Immunoprecipitation  IHC-P=Immunohistochemistry (Paraffin)  IHC-F=Immunohistochemistry (Frozen)  IF-IC=Immunofluorescence (Immunocytochemistry)  F=Flow Cytometry
Reactivity Key:  H=Human  M=Mouse  R=Rat  Mk=Monkey  C=Chicken  Mi=Mink  Pg=Pig  Sc=S. cerevisiae
Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Specificity / Sensitivity

Each antibody in the p70 S6 Kinase Substrates Antibody Sampler Kit detects endogenous levels of its target protein. p70 S6 Kinase (49D7) Antibody #2708 also recognizes p85 S6 Kinase. Phospho-p70 S6 Kinase (Thr389) (108D2) Rabbit mAb #9234 also detects p85 S6 Kinase when phosphorylated at Thr412 and possibly S6KII when phosphorylated at Thr401. The other antibodies in the kit do not cross react with other proteins.

Western Blotting

Western Blotting

Western blot analysis of extracts from PC12, NIH/3T3, and SK-N-MC cells, using p70 S6 Kinase (49D7) Rabbit mAb #2708.

Western Blotting

Western Blotting

Western blot analysis of extracts from HeLa cells using Phospho-eIF4B (Ser422) Antibody #3591 (upper) or eIF4B Antibody #3592 (lower). 48 hours following siRNA transfection, cells were treated with Rapamycin (50 nM) and U0126 (10 µM) as indicated.

Western Blotting

Western Blotting

Western blot analysis of extracts from SW-13 cells (starved for 18 hours) treated with calf intestinal alkaline phosphatase (CIP) or 20% fetal bovine serum for 30 minutes, and extracts from HeLa cells (starved for 18 hours) treated with 200 nM TPA for 30 minutes, using Phospho-eEF2k (Ser366) Antibody #3691 (upper) or eEF2k Antibody #3692 (lower).


Western Blotting

Western Blotting

Western blot analysis of extracts from PC12 and NIH/3T3 cells, treated with λ phosphatase, FBS or PDGF as indicated, using Phospho-S6 Ribosomal Protein (Ser235/236) (D57.2.2E) XP® Rabbit mAb #4858 (upper) or S6 Ribosomal Protein (5G10) Rabbit mAb #2217 (lower).

Western Blotting

Western Blotting

Western blot analysis of extracts from MCF7 and NIH/3T3 cells, treated with 100 nM insulin (10 min) or 20% FBS (30 min) as indicated, using Phospho-S6 Ribosomal Protein (Ser240/244) (D68F8) XP® Rabbit mAb #5364 (upper) or S6 Ribosomal Protein (5G10) Rabbit mAb #2217 (lower).

Western Blotting

Western Blotting

Western blot analysis of extracts from serum starved or serum treated (20%) 293, NIH/3T3, and PC12 cells, using Phospho-p70 S6 Kinase (Thr389) (108D2) Rabbit mAb #9234 (upper), or p70 S6 Kinase (49D7) rabbit mAb #2708 (lower).


Description

The p70 S6 Kinase Substrates Antibody Sampler Kit provides a fast and economical means of evaluating several substrates of p70 S6 Kinase. The kit contains enough primary and secondary antibody to perform four Western blot experiments.

Source / Purification

Antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding the amino-terminus of human p70 S6 Kinase; and to synthetic phosphopeptides corresponding to residues surrounding Thr389 of human p70 S6 Kinase; Ser235 and 236 of human ribosomal protein; Ser240 and 244 of human ribosomal protein; Ser422 of human eIF4B; and Ser366 of human eEF2k. Polyclonal antibodies are purified by protein A and peptide affinity chromatography.

Background

p70 S6 kinase is a mitogen activated Ser/Thr protein kinase that is required for cell growth and G1 cell cycle progression (1,2). p70 S6 kinase phosphorylates the S6 protein of the 40S ribosomal subunit and is involved in translational control of 5' oligopyrimidine tract mRNAs (1). Important S6 ribosomal protein phosphorylation sites include several residues (Ser235, Ser236, Ser240, Ser244) located wtihin a small, carboxy-terminal region of the S6 protein (3,4). p70 S6 kinase has been shown to phosphorylate eIF4B at the rapamycin-sensitive site Ser422 in vivo, and a Ser422Ala mutant of eIF4B shows diminished activity in an in vitro translation assay (5). Phosphorylation of eEF2K by p70 S6 kinase and p90RSK leads to inactivation of eEF2K (6), facilitating the dephosphorylation of eEF2 and thus promoting translation.

  1. Pullen, N. and Thomas, G. (1997) FEBS Lett 410, 78-82.
  2. Dufner, A. and Thomas, G. (1999) Exp Cell Res 253, 100-9.
  3. Ferrari, S. et al. (1991) J Biol Chem 266, 22770-5.
  4. Flotow, H. and Thomas, G. (1992) J Biol Chem 267, 3074-8.
  5. Raught, B. et al. (2004) EMBO J 23, 1761-9.
  6. Wang, X. et al. (2001) EMBO J 20, 4370-9.

Application References

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Protocols

Companion Products

Selected rabbit monoclonal antibodies are produced under license (granting certain rights including those under U. S. Patent No. 5,675,063 and/or U.S.S.N. 11/476,277) from Epitomics, Inc.

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

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