Cell Signaling Technology

Product Pathways - Metabolism

Phospho-Insulin/IGF Receptor Antibody Sampler Kit #8338

Kit Includes Quantity Applications Reactivity MW (kDa) Isotype
Phospho-IGF-I Receptor β (Tyr1135) (DA7A8) Rabbit mAb #3918 40 µl W H M R 95 Rabbit IgG
Phospho-IGF-I Receptor β (Tyr1131)/Insulin Receptor β (Tyr1146) Antibody #3021 40 µl W IP H M R (B) 95 Rabbit
Phospho-IGF-I Receptor β (Tyr1135/1136)/Insulin Receptor β (Tyr1150/1151) (19H7) Rabbit mAb #3024 40 µl W H M R (B) 95 Rabbit IgG
Phospho-IGF-I Receptor β (Tyr980) (C14A11) Rabbit mAb #4568 40 µl W H M R 95 Rabbit IgG
Insulin Receptor β (4B8) Rabbit mAb #3025 40 µl W IP H M R 95 Rabbit IgG
Anti-rabbit IgG, HRP-linked Antibody #7074 100 µl Goat
IGF-I Receptor β (D23H3) XP® Rabbit mAb #9750 40 µl W IP IF-IC F H M R Mk 95 Rabbit IgG

Applications Key:  W=Western Blotting  IP=Immunoprecipitation  IF-IC=Immunofluorescence (Immunocytochemistry)  F=Flow Cytometry
Reactivity Key:  H=Human  M=Mouse  R=Rat  Mk=Monkey  B=Bovine
Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Specificity / Sensitivity

Each antibody in the Phospho-Insulin/IGF Receptor Antibody Sampler Kit recognizes endogenous levels of the specific target protein, with activation state-specific antibodies recognizing target proteins only when phosphorylated at the indicated residues. IGF-I Receptor ß (D23H3) XP® Rabbit mAb detects endogenous levels of total IGF-I receptor ß protein. This antibody does not cross-react with insulin receptor.Phospho-IGF-I Receptor beta (Tyr1135) (DA7A8) Rabbit mAb detects endogenous levels of IGF-I receptor only when phosphorylated at Tyr1135. This antibody cross-reacts with Tyr1150 of insulin receptor and may also cross-react with other overexpressed related tyrosine-phosphorylated tyrosine kinases. Phospho-IGF-I Receptor ß (Tyr1131)/Insulin Receptor ß (Tyr1146) Antibody detects endogenous levels of Tyr1131-phosphorylated IGF-I receptor and Tyr1146-phosphorylated insulin receptor. This antibody cross-reacts with activated PDGF, FGF and EGF receptors; ErbB2; and c-Met. Phospho-IGF-I Receptor ß (Tyr1135/1136)/Insulin Receptor ß (Tyr1150/1151) (19H7) Rabbit mAb detects endogenous levels of IGF-I receptor and insulin receptor only when phosphorylated at tyrosine 1135/1136 or tyrosine 1150/1151, and does not cross-react with other related tyrosine-phosphorylated tyrosine kinases. Phospho-IGF-I Receptor ß (Tyr980) (C14A11) Rabbit mAb detects endogenous levels of IGF-I receptor ß protein only when phosphorylated at Tyr980, and may cross-react with activated insulin receptors and FLT3. Insulin Receptor ß (4B8) Rabbit mAb detects endogenous levels of total insulin receptor ß and does not cross-react with IGF-I receptor ß.

Western Blotting

Western Blotting

Western blot analysis of extracts from 3T3-L1 adipocytes, untreated or insulin-treated (100 nM for the indicated times), using Phospho-IGF-I Receptor β (Tyr1131)/Insulin Receptor β (Tyr1146) Antibody #3021 (upper) or control IR antibody (lower).

Western Blotting

Western Blotting

Western blot analysis of extracts from HeLa cells, untreated (-) or treated with IGF and serum starved (+), as well as H-4-II-E cells, untreated (-) or treated with insulin and serum starved (+), using Phospho-IGF-I Receptor β (Tyr1135/1136)/Insulin Receptor β (Tyr1150/1151) (19H7) Rabbit mAb #3024.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using Insulin Receptor β (4B8) Rabbit mAb #3025.


Western Blotting

Western Blotting

Western blot analysis of extracts from MCF-7 cells, untreated (-) or treated with IGF-I (+), using Phospho-IGF-I Receptor β (Tyr1135) (DA7A8) Rabbit mAb #3918 (upper) and IGF-I Receptor β Antibody #3027 (lower).

Western Blotting

Western Blotting

Western blot analysis of extracts from MCF7 cells, untreated (-) or treated with IGF-I (+), using Phospho-IGF-I Receptor β (Tyr980) (C14A11) Rabbit mAb #4568 (upper) and IGF-I Receptor β (111A9) Rabbit mAb #3018 (lower).

Western Blotting

Western Blotting

Western blot analysis of extracts from 293 (IGF-I receptor β+) and SK-UT-1 (IGF-I receptor β-) cells using IGF-I Receptor β (D23H3) XP® Rabbit mAb #9750 (upper) or β-Actin Antibody #4967 (lower).


Description

The Phospho-Insulin/IGF Receptor Antibody Sampler Kit provides an economical means of evaluating total Insulin Receptor and IGF-I Receptor β protein levels as well as Insulin and IGF-I Receptor β phosphorylated at specific sites. The kit includes enough antibody to perform four western blot experiments with each primary antibody.

Source / Purification

Monoclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to the carboxy-terminal residues of human IGF-IR β or residues surrounding Tyr960 of human insulin receptor β. Activation state-specific monoclonal antibodies are produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Tyr1135 of human IGF-I Receptor β, Tyr1135/1136 of human IGF-I Receptor β, or Tyr980 of human IGF-I Receptor β. Activation state-specific polyclonal antibodies are produced by immunizing animals with a synthetic phosphopeptide corresponding to residues of human IGF-I Receptor β. Polyclonal antibodies are purified by protein A and peptide affinity chromatography.

Background

Type I insulin-like growth factor receptor (IGF-IR) is a transmembrane receptor tyrosine kinase that is widely expressed in many cell lines and cell types within fetal and postnatal tissues (1-3). Receptor autophosphorylation follows binding of the IGF-I and IGF-II ligands. Three tyrosine residues within the kinase domain (Tyr1131, Tyr1135, and Tyr1136) are the earliest major autophosphorylation sites (4). Phosphorylation of these three tyrosine residues is necessary for kinase activation (5,6). Insulin receptors (IRs) share significant structural and functional similarity with IGF-I receptors, including the presence of an equivalent tyrosine cluster (Tyr1146/1150/1151) within the kinase domain activation loop. Tyrosine autophosphorylation of IRs is one of the earliest cellular responses to insulin stimulation (7). Autophosphorylation begins with phosphorylation at Tyr1146 and either Tyr1150 or Tyr1151, while full kinase activation requires triple tyrosine phosphorylation (8).

  1. Adams, T.E. et al. (2000) Cell. Mol. Life Sci. 57, 1050-1093.
  2. Baserga, R. et al. (2000) Oncogene 19, 5574-5581.
  3. Scheidegger, K.J. et al. (2000) J. Biol. Chem. 275, 38921-38928.
  4. Hernandez-Sanchez, C. et al. (1995) J. Biol. Chem. 270, 29176-29181.
  5. Lopaczynski, W. et al. (2000) Biochem. Biophys. Res. Commun. 279, 955-960.
  6. Baserga, R. et al. (1999) Exp. Cell Res. 253, 1-6.
  7. White, M.F. et al. (1985) J. Biol. Chem. 260, 9470-9478.
  8. White, M.F. et al. (1988) J. Biol. Chem. 263, 2969-2980.

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