Product Pathways - Translational Control
4E-BP Antibody Sampler Kit #9955
|Kit Includes||Quantity||Applications||Reactivity||MW (kDa)||Isotype|
|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|
|Non-phospho-4E-BP1 (Thr46) (87D12) Rabbit mAb #4923||40 µl||W F||H M R Mk||15-20||Rabbit IgG|
|Phospho-4E-BP1 (Ser65) Antibody #9451||40 µl||W IP||H M R Mk||15 to 20||Rabbit|
|4E-BP1 (53H11) Rabbit mAb #9644||40 µl||W IP IHC-P IF-IC F||H M R Mk||15-20||Rabbit IgG|
|4E-BP2 Antibody #2845||40 µl||W IP IHC-P F||H M R Mk B||15 to 20||Rabbit|
|Phospho-4E-BP1 (Thr70) Antibody #9455||40 µl||W IP||H M R Mk||15 to 20||Rabbit|
|Anti-rabbit IgG, HRP-linked Antibody #7074||100 µl||Goat|
Reactivity Key: H=Human M=Mouse R=Rat Mk=Monkey Dm=D. melanogaster B=Bovine
Species enclosed in parentheses are predicted to react based on 100% sequence homology.
Specificity / Sensitivity
Phospho-4E-BP1 (Thr37/46) Rabbit mAb detects endogenous levels of 4E-BP1 only when phosphorylated at Thr37 and/or Thr46, and may cross-react with 4E-BP2 and 4E-BP3 when phosphorylated at equivalent sites. Nonphospho-4E-BP1 (Thr46) (87D12) Rabbit mAb detects endogenous levels of 4E-BP1 only when dephosphorylated at Thr46. This antibody cross-reacts with 4E-BP2 and 4E-BP3 dephosphorylated at equivalent sites. Phospho-4E-BP1 (Ser65) Antibody detects endogenous levels of 4E-BP1 when phosphorylated at Ser65, and may also recognize 4E-BP1 when phosphorylated at Ser101. Phospho-4E-BP1 (Ser65) (174A9) Rabbit mAb detects endogenous levels of 4E-BP1 when phosphorylated at Ser65. Phospho-4E-BP1 (Thr70) Antibody detects endogenous levels of 4E-BP1 only when phosphorylated at Thr70. 4E-BP1 (53H11) Rabbit mAb detects endogenous levels of total 4E-BP1 protein. 4E-BP2 Antibody detects endogenous levels of total 4E-BP2 independent of phosphorylation and does not cross-react significantly with 4E-BP1.
Western blot analysis of bacterially expressed GST-4E-BP1 and of extracts from NIH/3T3 cells using 4E-BP2 Antibody #2845 and 4E-BP1 Antibody #9452.
Western blot analysis of extracts from 293 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 depravation. Amino acids were then added back for 1 hour, and cells were either untreated (-) or treated with 100 nM insulin (+) for 30 minutes.
Western blot analysis of extracts from COS cells, λ phosphatase-treated or 20% FBS-treated, using Nonphospho-4E-BP1 (Thr46) (87D12) Rabbit mAb #4923 (upper), Phospho-4E-BP1 (Thr37/46) Antibody #9459 (middle), and 4E-BP1 Antibody #9452 (lower).
Western blot analysis of extracts from C6 and MEF cells using Phospho-4E-BP1 (Ser65) Antibody #9451 (upper) or 4E-BP1 (53H11) Rabbit mAb #9644 (lower).
The 4E-BP Antibody Sampler Kit provides an economical means to investigate regulation of cap-dependent translation within the cell. The kit contains primary and secondary antibodies to perform four Western blots with each antibody.
Source / Purification
Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Thr37 and Thr46 of mouse 4E-BP1, residues surrounding Thr46 of human 4E-BP1, or Ser112 of human 4E-BP1. Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to the residues at the carboxy-terminus of human 4E-BP2 (#2845), or phosphopeptides surrounding mouse Ser65 (#9451) and human Thr70 (#5078) 4E-BP1. Polyclonal antibodies were purified by protein A and peptide affinity chromatography.
Translation repressor protein 4E-BP1 (also known as PHAS-1) inhibits cap-dependent translation by binding to the translation initiation factor eIF4E. Hyperphosphorylation of 4E-BP1 disrupts this interaction and results in activation of cap-dependent translation (1). Both the PI3 kinase/Akt pathway and FRAP/mTOR kinase regulate 4E-BP1 activity (2,3). Multiple 4E-BP1 residues are phosphorylated in vivo (4). While phosphorylation by FRAP/mTOR at Thr37 and Thr46 does not prevent the binding of 4E-BP1 to eIF4E, it is thought to prime 4E-BP1 for subsequent phosphorylation at Ser65 and Thr70 (5).
4E-BP2 and 4E-BP3 share high sequence homology with 4E-BP1, including conservation of the major FRAP/mTOR-dependent phosphorylation sites. Preliminary data suggests that phosphorylation of 4E-BP2 is regulated in a similar manner to that of 4E-BP1, although phosphorylation of this protein has not been as extensively studied (6).
- Pause, A. et al. (1994) Nature 371, 762-767.
- Brunn, G.J. et al. (1997) Science 277, 99-101.
- Gingras, A.C. et al. (1998) Genes Dev. 12, 502-513.
- Fadden, P. et al. (1997) J. Biol. Chem. 272, 10240-10247.
- Gingras, A.C. et al. (1999) Genes Dev. 13, 1422-1437.
- Lin, T.A. and Lawrence, J.C. (1996) J. Biol. Chem. 271, 30199-30204.
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- 9459 Phospho-4E-BP1 (Thr37/46) Antibody
- 9452 4E-BP1 Antibody
- 1052 Phospho-4E-BP1 (Thr37/46) Blocking Peptide
- 9456 Phospho-4E-BP1 (Ser65) (174A9) Rabbit mAb
- 7216 PathScan® Phospho-4E-BP1 (Thr37/Thr46) Sandwich ELISA Kit
- 2441 Phospho-eIF4G (Ser1108) Antibody
- 2498 eIF4G Antibody
- 9741 Phospho-eIF4E (Ser209) Antibody
- 9742 eIF4E Antibody
- 7071 Phototope®-HRP Western Blot Detection System, 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
- 7074 Anti-rabbit IgG, HRP-linked Antibody
For Research Use Only. Not For Use In Diagnostic Procedures.