Cell Signaling Technology

Product Pathways - Translational Control

Phospho-S6 Ribosomal Protein (Ser235/236) (2F9) Rabbit mAb (Alexa Fluor® 488 Conjugate) #4854

Applications Reactivity Sensitivity Isotype
IF-F IF-IC F H M R Endogenous Rabbit IgG

Applications Key:  IF-F=Immunofluorescence (Frozen)  IF-IC=Immunofluorescence (Immunocytochemistry)  F=Flow Cytometry
Reactivity Key:  H=Human  M=Mouse  R=Rat
Species cross-reactivity is determined by western blot. Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Protocols

Specificity / Sensitivity

Phospho-S6 Ribosomal Protein (Ser235/236) (2F9) Rabbit mAb detects endogenous levels of ribosomal protein S6 only when phosphorylated at serine 235 and 236.

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Ser235 and Ser236 of human ribosomal protein S6. The antibody was conjugated to Alexa Fluor® 488 under optimal conditions with an F/P ratio of 2-6.

Flow Cytometry

Flow Cytometry

Flow cytometric analysis of Jurkat cells, untreated (green), or LY294002, Wortmannin and U0126-treated (blue), using Phospho-S6 Ribosomal Protein (Ser235/236) (2F9) Rabbit mAb (Alexa Fluor® 488 Conjugate) (#4854).

IF-IC

IF-IC

Confocal immunofluorescent analysis of two Gefinitib (Iressa) treated non-small cell lung cancer cell lines. HCC827 cells have the E746_A750 deletion in exon 19 of the EGFR gene, and are highly sensitive to Gefitinib. H1975 cells have the (T790M) mutation that confers Gefitinib-resistance. Both cell lines were treated and then double-labeled with Phospho-S6 Ribosomal Protein Rabbit mAb (Alexa Fluor® 488 Conjugate) #4854 and Phospho-Tyrosine Mouse mAb (Alexa Fluor® 647 Conjugate) #9415. Untreated HCC827 (A) and H1975 (C) cells show bright phospho-S6 (green) and phospho-tyrosine (red pseudocolor) label. Phospho-S6 and phospho-tyrosine signals dramatically decrease following Gefitinib treatment in HCC827 cells (B), with little or no change in the Gefitinib-resistant H1975 cells (D).

IF-F

IF-F

Confocal immunofluorescent analysis of rat brain showing the hippocampus (left) and striatum (right) labeled with Phospho-S6 Ribosomal Protein (Ser235/236) (2F9) Rabbit mAb (Alexa Fluor® 488 Conjugate) (green), EGR1 Antibody (red) #4152, and Phospho-Histone H3 (Ser10) (6G3) Mouse mAb #9706 (blue) following ischemia with 30 minute reperfusion.


Description

This Cell Signaling Technology antibody is conjugated to Alexa Fluor® 488 fluorescent dye and tested in-house for direct flow cytometry and immunofluorescent analysis in human cells. The antibody is expected to exhibit the same species cross-reactivity as the unconjugated Phospho-S6 Ribosomal Protein (Ser235/236) (2F9) Rabbit mAb #4856.

Background

One way that growth factors and mitogens effectively promote sustained cell growth and proliferation is by upregulating mRNA translation (1,2). Growth factors and mitogens induce the activation of p70 S6 kinase and the subsequent phosphorylation of the S6 ribosomal protein. Phosphorylation of S6 ribosomal protein correlates with an increase in translation of mRNA transcripts that contain an oligopyrimidine tract in their 5' untranslated regions (2). These particular mRNA transcripts (5'TOP) encode proteins involved in cell cycle progression, as well as ribosomal proteins and elongation factors necessary for translation (2,3). Important S6 ribosomal protein phosphorylation sites include several residues (Ser235, Ser236, Ser240, and Ser244) located within a small, carboxy-terminal region of the S6 protein (4,5).

  1. Dufner, A. and Thomas, G. (1999) Exp. Cell Res. 253, 100-109.
  2. Peterson, R.T. and Schreiber, S.L. (1998) Curr. Biol. 8, R248-R250.
  3. Jefferies, H.B. et al. (1997) EMBO J. 16, 3693-3704.
  4. Ferrari, S. et al. (1991) J. Biol. Chem. 266, 22770-22775.
  5. Flotow, H. and Thomas, G. (1992) J. Biol. Chem. 267, 3074-3078.

Application References

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Companion Products

Rabbit monoclonal antibody is produced under license (granting certain rights including those under U. S. Patent No. 5,675,063 and U.S.S.N. 11/476,277) from Epitomics, Inc.The Alexa Fluor® dye antibody conjugates in this product are sold under license from Molecular Probes, Inc., for research use only, except for use in combination with DNA microarrays. The Alexa Fluor® dyes (except for Alexa Fluor® 430 dye) are covered by pending and issued patents.Alexa Fluor® is a registered trademark of Molecular Probes, Inc.

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

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