Revision 6

#9644Store at -20C

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, W-S, IP, IHC-P, IF-IC, FC-FP

REACTIVITY:

H M R Mk

SENSITIVITY:

Endogenous

MW (kDa):

15-20

Source/Isotype:

Rabbit IgG

UniProt ID:

#Q13541

Entrez-Gene Id:

1978

Product Information

Product Usage Information

Application Dilution
Western Blotting 1:1000
Simple Western™ 1:50 - 1:250
Immunoprecipitation 1:50
Immunohistochemistry (Paraffin) 1:1200 - 1:4800
Immunofluorescence (Immunocytochemistry) 1:800 - 1:3200
Flow Cytometry (Fixed/Permeabilized) 1:1600

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.

For a carrier free (BSA and azide free) version of this product see product #42235.

Specificity / Sensitivity

4E-BP1 (53H11) Rabbit mAb detects endogenous levels of total 4E-BP1 protein.

Species Reactivity:

Human, Mouse, Rat, Monkey

Source / Purification

4E-BP1 (53H11) Rabbit mAb is produced by immunizing rabbits with a synthetic peptide corresponding to residues surrounding Ser112 of human 4E-BP1.

Background

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).

  1. Pause, A. et al. (1994) Nature 371, 762-7.
  2. Brunn, G.J. et al. (1997) Science 277, 99-101.
  3. Gingras, A.C. et al. (1998) Genes Dev 12, 502-13.
  4. Fadden, P. et al. (1997) J Biol Chem 272, 10240-7.
  5. Gingras, A.C. et al. (1999) Genes Dev 13, 1422-37.

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 W-S: Simple Western™ IP: Immunoprecipitation IHC-P: Immunohistochemistry (Paraffin) IF-IC: Immunofluorescence (Immunocytochemistry) FC-FP: Flow Cytometry (Fixed/Permeabilized)

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.
Alexa Fluor is a registered trademark of Life Technologies Corporation.
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 6
#9644

4E-BP1 (53H11) Rabbit mAb

Western Blotting Image 1: 4E-BP1 (53H11) Rabbit mAb Expand Image
Western blot analysis of extracts from control HeLa cells (Lane 1) or HeLa cells with a targeted mutation in the gene encoding 4E-BP1 (Lane 2) using 4E-BP1 (53H11) Rabbit mAb (upper) or β-actin (13E5) Rabbit mAb #4970 (lower). The change in 4E-BP1 molecular weight in the mutated HeLa cells confirms the specificity of the antibody for 4E-BP1.
Western Blotting Image 2: 4E-BP1 (53H11) Rabbit mAb Expand Image
Western blot analysis of extracts from various cell lines using 4E-BP1 (53H11) Rabbit mAb.
Western Blotting Image 1: 4E-BP1 (53H11) Rabbit mAb Expand Image
Simple Western™ analysis of lysates (0.1 mg/mL) from HepG2 cells using 4E-BP1 (53H11) Rabbit mAb #9644. The virtual lane view (left) shows the target band (as indicated) at 1:50 and 1:250 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:50 (blue line) and 1:250 (green line) dilutions of primary antibody. This experiment was performed under reducing conditions on the Jess™ Simple Western instrument from ProteinSimple, a BioTechne brand, using the 2-40 kDa separation module.
No image available
Immunohistochemistry Image 1: 4E-BP1 (53H11) Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human breast carcinoma using 4E-BP1 (53H11) Rabbit mAb in the presence of control peptide (left) or 4E-BP1 blocking peptide #1053 (right).
Immunohistochemistry Image 2: 4E-BP1 (53H11) Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human hepatocellular carcinoma using 4E-BP1 (53H11) Rabbit mAb.
Immunohistochemistry Image 3: 4E-BP1 (53H11) Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human lung carcinoma using 4E-BP1 (53H11) Rabbit mAb.
Immunohistochemistry Image 4: 4E-BP1 (53H11) Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded mouse spleen, 4E-BP1/2 wild type (left) or 4E-BP1 knockout (right), using 4E-BP1 (53H11) Rabbit mAb. 4E-BP wild type and knockout tissues kindly provided by Dr. Nahum Sonenberg, McGill University.
Immunofluorescence Image 1: 4E-BP1 (53H11) Rabbit mAb Expand Image
Confocal immunofluorescent analysis of HeLa cells using 4E-BP1 (53H11) Rabbit mAb (green). Actin filaments have been labeled with Alexa Fluor® 555 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
Immunofluorescence Image 2: 4E-BP1 (53H11) Rabbit mAb Expand Image
Confocal immunofluorescent analysis of 293 cells, expressing either nontargeting shRNA (left) or shRNA targeting 4E-BP1/2 (right), using 4E-BP1 (53H11) Rabbit mAb #9644 (green). Red = Propidium Iodide (PI)/RNase Staining Solution #4084.
Flow Cytometry Image 1: 4E-BP1 (53H11) Rabbit mAb Expand Image
Flow cytometric analysis of 293 cells, transfected with control shRNA (green) or 4E-BP1-specific shRNA (blue) using 4E-BP1 (53H11) Rabbit mAb (solid lines) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')₂ Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.