Product Pathways - Translational Control
Translational Control Antibody Sampler Kit #9918
|Kit Includes||Quantity||Applications||Reactivity||MW (kDa)||Isotype|
|Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb #4060||40 µl||W IP IHC-P IHC-F IF-IC F||H M R Hm Mk Dm Z B (C) (X) (Dg) (Pg)||60||Rabbit IgG|
|Phospho-p70 S6 Kinase (Thr389) (108D2) Rabbit mAb #9234||40 µl||W||H M R Mk (C)||70, 85||Rabbit IgG|
|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-eIF2α (Ser51) (D9G8) XP® Rabbit mAb #3398||40 µl||W IP IHC-P||H M R Mk Dm||38||Rabbit IgG|
|Phospho-4E-BP1 (Thr37/46) (236B4) Rabbit mAb #2855||40 µl||W IHC-P IF-IC F||H M R Mk Dm||15 to 20||Rabbit IgG|
|Phospho-eIF4E (Ser209) Antibody #9741||40 µl||W||H M R Mk||25||Rabbit|
|Rapamycin #9904||2 micrograms||914.2|
|LY294002 #9901||0.3 milligrams||307.34|
|Anti-rabbit IgG, HRP-linked Antibody #7074||100 µl||Goat|
Reactivity Key: H=Human M=Mouse R=Rat Hm=Hamster Mk=Monkey C=Chicken Mi=Mink Dm=D. melanogaster X=Xenopus Z=Zebrafish B=Bovine Dg=Dog Pg=Pig Sc=S. cerevisiae
Species enclosed in parentheses are predicted to react based on 100% sequence homology.
Specificity / Sensitivity
Each phospho-specific antibody in the Translational Control Antibody Sampler Kit detects the intended target only when phosphorylated at the indicated site. Phospho-4E-BP1 (Thr37/46) (236B4) Rabbit mAb may cross-react with 4E-BP2 and 4E-BP3 when phosphorylated at equivalent sites.
Western blot analysis of extracts from 293T cells using 4E-BP1 Antibody #9452 (upper) and Phospho-4E-BP1 (Thr37/46) Antibody #2855 (lower). The cells were starved for 24 hours in serum-free medium and underwent a 1 hour amino acid deprivation. Amino acids were replenished for 1 hour. Cells were then either untreated (-) or treated with 100 nM insulin (+) for 30 minutes.
Western blot analysis of extracts from C2C12 cells, untreated or thapsigargin-treated, using Phospho-eIF2α (Ser51) (D9G8) XP® Rabbit mAb #3398 (upper) or eIF2α Antibody #9722 (lower).
Western blot analysis of extracts from PC-3 cells, untreated or LY294002/wortmannin-treated, and NIH/3T3 cells, serum-starved or PDGF-treated, using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb #4060 (upper) or Akt (pan) (C67E7) Rabbit mAb #4691 (lower).
Western blot analysis of extracts from PC12 and NIH/3T3 cells, treated with λ phosphatase, 20% FBS (20 min) or 100 ng/ml PDGF (20 min) 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 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).
Western blot analysis of extracts from NIH/3T3 cells, untreated or treated with serum, PD98059 or Dexamethasone, using Phospho-eIF4E (Ser209) Antibody #9741 (upper) or eIF4E Antibody #9742 (lower).
The Translational Control Antibody Sampler Kit provides a fast and economical means of evaluating multiple proteins involved in translational control. The kit contains enough primary and secondary antibody to perform four Western blot experiments, as well as specific inhibitors of PI3 kinase and mTOR/FRAP.
Source / Purification
Phospho-specific polyclonal antibodies are produced by immunizing animals with synthetic phosphopeptides corresponding to residues surrounding Ser209 of human elF4E. Polyclonal antibodies are purified by protein A and peptide affinity chromatography. Phospho-specific rabbit monoclonal antibodies are produced by immunizing animals with synthetic phosphopeptides corresponding to residues surrounding Ser473 of mouse Akt, Thr37 and Thr46 of mouse 4E-BP1, Ser51 of human eIF2α, Ser235 and Ser236 of human ribosomal protein S6, and Thr389 of human p70 S6 kinase.
Key steps in translational control occur at the level of eukaryotic initiation factor 4F (eIF4F) and p70 S6 kinase regulation. eIF4F is a complex whose functions include the recognition of the mRNA 5' cap structure. Several stimuli, such as insulin and various growth and survival factors, regulate the eIF4F complex and p70 S6 kinase primarily by triggering a signaling cascade dependent on sequential activation of PI3K, Akt/PKB and mTOR/FRAP kinases. Akt is activated by phosphorylation within the C-terminus at Ser473 and within the activation loop at Thr308 by phospholipid-dependent kinases. Inactivation in vivo of PI3K by the highly selective inhibitor LY294002 inhibits Akt and downstream elements of this cascade. Direct phosphorylation of mTOR/FRAP at Ser2448 by Akt is a key regulatory event controlling its kinase activity. mTOR/FRAP activity can be effectively blocked by Rapamycin, leading to inactivation of eukaryotic initiation factor 4E binding protein 1 (4E-BP1), an inhibitor of translation initiation, and activation of p70 S6 kinases. Inactivation of 4E-BP1 by sequential phosphorylation causes the release of eIF4E, which, together with eIF4G and other factors, forms a functional eIF4F cap binding complex. p70 S6 kinases phosphorylates the 40S ribosomal subunit protein S6 and stimulates the translation of 5' oligopyrimidine tract containing mRNAs. The Erk pathway is also involved in regulation at this level by regulating the eIF4E kinase, Mnk1, and activating p70 S6 kinase. Tuberin, a product of the tumor supressor gene TSG2, is directly phosphorylated atThr1462 by Akt/PKB. Tuberin inhibits the mammalian target of rapamycin, mTOR, which results in inhibition of p70 S6 kinase and activation of 4E-BP1 and, therefore, inhibition of translation.
- Gingras, A. et al. (1999) Annu. Rev. Biochem. 68, 913-963.
- Gingras, A. C. et al. (2001) Genes and Develop. 15, 807-826.
- Dennis, P. B. et al. (1999) Curr. Opin. Genet. Dev. 9, 49-54.
- Volarevic, S. and Thomas, G. (2000) Prog. Nucleic Acid Res. Mol. Biol. 65, 101-127.
- Pyronnet, S. et al. (2000) Biochem. Pharmacol. 60, 1237-1243.
- Dever, T.E. (2002) Cell 108, 545-556.
- Goncharova, E. et al. (2002) J. Biol. Chem. 277, 30958-30967.
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* Product-specific protocol.
- 2067 eIF4E (C46H6) Rabbit mAb
- 9742 eIF4E Antibody
- 9205 Phospho-p70 S6 Kinase (Thr389) Antibody
- 9206 Phospho-p70 S6 Kinase (Thr389) (1A5) Mouse mAb
- 9721 Phospho-eIF2α (Ser51) Antibody
- 2103 eIF2α (L57A5) Mouse mAb
- 9722 eIF2α Antibody
- 4051 Phospho-Akt (Ser473) (587F11) Mouse mAb
- 9271 Phospho-Akt (Ser473) Antibody
- 4691 Akt (pan) (C67E7) Rabbit mAb
- 9272 Akt Antibody
- 9644 4E-BP1 (53H11) Rabbit mAb
- 9451 Phospho-4E-BP1 (Ser65) Antibody
- 2215 Phospho-S6 Ribosomal Protein (Ser240/244) Antibody
- 2217 S6 Ribosomal Protein (5G10) Rabbit mAb
- 2317 S6 Ribosomal Protein (54D2) Mouse mAb
- 9202 p70 S6 Kinase Antibody
- 9456 Phospho-4E-BP1 (Ser65) (174A9) Rabbit mAb
- 7074 Anti-rabbit IgG, HRP-linked Antibody
- 7720 Prestained Protein Marker, Broad Range (Premixed Format)
- 7727 Biotinylated Protein Ladder Detection Pack
- 7003 20X LumiGLO® Reagent and 20X Peroxide
Selected rabbit monoclonal antibodies are produced under license (granting certain rights including those under U. S. Patent No. 5,675,063 and/orU.S.S.N. 11/476,277) from Epitomics, Inc.
For Research Use Only. Not For Use In Diagnostic Procedures.