anti igf 1r ab  (Cell Signaling Technology Inc)

Journal: Journal of the Endocrine Society

Article Title: Insulin receptor expression and its association with hyperinsulinemia in triple negative breast cancer

doi: 10.1210/jendso/bvag041

Figure Lengend Snippet: Representative images of IHC staining. IHC staining of TNBC specimens was performed as described in the Methods section with antibodies to IR, IGF-1R, pErk1/2, and FOXO3a. The images are shown in 40× magnification. Abbreviations: IGF-1R, IGR-1 receptor; IHC, immunohistochemistry; IR, insulin receptor; pErk1/2, phosphorylated Erk1/2; TNBC, triple negative breast cancer.

Article Snippet: Primary antibodies were anti-IR Ab (ab137747, Abcam, Cambridge, MA, RRID:AB_3717506) 1:300 dilution, anti-IGF-1R Ab (#3027, Cell Signaling, Danvers, CA, RRID:AB_2122378) 1:200 dilution, anti-FOXO3a Ab (#12829, Cell Signaling, RRID:AB_2636990) 1:3200 dilution, and anti-pErk1/2 (Thr202/Tyr204) Ab (#4370, Cell Signaling, RRID:AB_2315112) 1:400 dilution.

Techniques: Immunohistochemistry

Journal: Journal of the Endocrine Society

Article Title: Insulin receptor expression and its association with hyperinsulinemia in triple negative breast cancer

doi: 10.1210/jendso/bvag041

Figure Lengend Snippet: Representative images of protein localization for IR, IGF1R, FOXO3a, and pErk1/2. IHC staining of TNBC specimens was performed and evaluated for localization of proteins. The images are shown in 40× magnification. Abbreviations: IGF-1R, IGR-1 receptor; IHC, immunohistochemistry; IR, insulin receptor; pErk1/2, phosphorylated Erk1/2; TNBC, triple negative breast cancer.

Article Snippet: Primary antibodies were anti-IR Ab (ab137747, Abcam, Cambridge, MA, RRID:AB_3717506) 1:300 dilution, anti-IGF-1R Ab (#3027, Cell Signaling, Danvers, CA, RRID:AB_2122378) 1:200 dilution, anti-FOXO3a Ab (#12829, Cell Signaling, RRID:AB_2636990) 1:3200 dilution, and anti-pErk1/2 (Thr202/Tyr204) Ab (#4370, Cell Signaling, RRID:AB_2315112) 1:400 dilution.

Techniques: Immunohistochemistry

Journal: Journal of the Endocrine Society

Article Title: Insulin receptor expression and its association with hyperinsulinemia in triple negative breast cancer

doi: 10.1210/jendso/bvag041

Figure Lengend Snippet: Overall survival for women with breast cancer based on IR and IGF-1R protein expression in KMPlot.com . KMPlot.com was used to generate overall survival Kaplan-Meier plots. The dataset used was not from our cohort but from Tang and colleagues ; n = 65 patients had IR (A) and IGF1R (B) protein expression available. The patients were split using the “auto select best cutoff” percentile option in KMPlot. No other restrictions were added to the analysis. The plots show hazard ratios with 95% confidence intervals in parentheses . Abbreviations: IGF-1R, IGR-1 receptor; IR, insulin receptor.

Article Snippet: Primary antibodies were anti-IR Ab (ab137747, Abcam, Cambridge, MA, RRID:AB_3717506) 1:300 dilution, anti-IGF-1R Ab (#3027, Cell Signaling, Danvers, CA, RRID:AB_2122378) 1:200 dilution, anti-FOXO3a Ab (#12829, Cell Signaling, RRID:AB_2636990) 1:3200 dilution, and anti-pErk1/2 (Thr202/Tyr204) Ab (#4370, Cell Signaling, RRID:AB_2315112) 1:400 dilution.

Techniques: Expressing

Journal: Frontiers in Endocrinology

Article Title: Maternal Diabetes Impairs Insulin and IGF-1 Receptor Expression and Signaling in Human Placenta

doi: 10.3389/fendo.2021.621680

Figure Lengend Snippet: IR and IGF-1R content and phosphorylation in placental tissues. Insulin receptor protein content was detected by immunoblotting with a polyclonal anti-insulin receptor β-subunit. (A) The protein content of IGF-1R in placenta samples was detected by immunoblotting using a polyclonal anti-IGF-1R β-subunit. (B) Solubilized placental tissues were subjected to immunoprecipitation with an anti-IR or anti IGF-1R. The resulting immune complexes were resolved by SDS-PAGE and immunoblotting with an anti-PY antibody to detect receptor phosphorylation (A, B) . Protein content was normalized against β-actin, whereas phosphorylated proteins were normalized against their total amount. Each bar represents mean ± SE of all placental samples (n=40 for NGT, n=20 for GDM and n=20 for T1D). A representative Western-blot is included in the bottom (five different patients for each group). IR-A mRNA expression in placental tissue measured by RT-PCR. The bar graphs show the quantitation of IR-A mRNA (mean ± SE) in all samples for each group (C) . *p < 0.05; **p < 0.001, after adjusting, in multivariate logistic regression models, for age, pre-pregnancy BMI, gestational age at delivery and type of delivery.

Article Snippet: For immunoprecipitation studies, equal amounts of placental extracts (1 mg) were subjected to immunoprecipitation between protein G-Sepharose beads and anti-insulin receptor NB400-142 (MA-20) from Novus Biologicals (Littleton, USA) or anti-IGF-1R (Ab-1) mouse mAb (αIR3) from Calbiochem (San Diego, CA) for 2 h at 4°C, washed three times with lysis buffer, then eluted with Laemmli buffer 2 x for 5 min at 100°C.

Techniques: Phospho-proteomics, Western Blot, Immunoprecipitation, SDS Page, Expressing, Reverse Transcription Polymerase Chain Reaction, Quantitation Assay

Journal: Frontiers in Endocrinology

Article Title: Maternal Diabetes Impairs Insulin and IGF-1 Receptor Expression and Signaling in Human Placenta

doi: 10.3389/fendo.2021.621680

Figure Lengend Snippet: IR and IGF-1R content and activation in relationship to the maternal capillary blood glucose level during labor and delivery. IR content and phosphorylation (A) , and IGF-1R content and phosphorylation (B) in placental tissues of patients with peripartum blood glucose <90 mg/dl (N=25) or ≥90 mg/dl (N=15). Each bar represents mean ± SE of the samples indicated above. ***p < 0.0001. A representative Western-blot is included in the bottom (five different patients for each group). IR-A mRNA expression in placental tissues of patients with peripartum average blood glucose <90 mg/dl (N=25) or ≥90 mg/dl (N=15) (C) and of patients with peri-partum average blood glucose <120 mg/dl (N=30) or ≥120 mg/dl (N=10) (D) . Each bar represents mean ± SE of the samples indicated above. *p < 0.05.

Article Snippet: For immunoprecipitation studies, equal amounts of placental extracts (1 mg) were subjected to immunoprecipitation between protein G-Sepharose beads and anti-insulin receptor NB400-142 (MA-20) from Novus Biologicals (Littleton, USA) or anti-IGF-1R (Ab-1) mouse mAb (αIR3) from Calbiochem (San Diego, CA) for 2 h at 4°C, washed three times with lysis buffer, then eluted with Laemmli buffer 2 x for 5 min at 100°C.

Techniques: Activation Assay, Phospho-proteomics, Western Blot, Expressing

Journal: PLoS ONE

Article Title: Evaluation of a Novel Hexavalent Humanized Anti-IGF-1R Antibody and Its Bivalent Parental IgG in Diverse Cancer Cell Lines

doi: 10.1371/journal.pone.0044235

Figure Lengend Snippet: Epitope analysis by cross-blocking with anti-IGF-1R antibodies of known epitopes.

Article Snippet: Additional anti-human IGF-1R antibodies were obtained from Calbiochem for Ab-1 (clone αIR3), Ab-3 (Clone 33255.111), and Ab-4 (clone 24–31); from Santa Cruz Biotechnology for 2C8, 1H7, 3B7, and C-20; from Millipore for 24–57 and 24–31; and from Invitrogen for 17–69, 24–60, and 1–2.

Techniques:

Journal: PLoS ONE

Article Title: Evaluation of a Novel Hexavalent Humanized Anti-IGF-1R Antibody and Its Bivalent Parental IgG in Diverse Cancer Cell Lines

doi: 10.1371/journal.pone.0044235

Figure Lengend Snippet: Epitope analysis by cross-blocking with anti-IGF-1R antibodies of known epitopes (continued).

Article Snippet: Additional anti-human IGF-1R antibodies were obtained from Calbiochem for Ab-1 (clone αIR3), Ab-3 (Clone 33255.111), and Ab-4 (clone 24–31); from Santa Cruz Biotechnology for 2C8, 1H7, 3B7, and C-20; from Millipore for 24–57 and 24–31; and from Invitrogen for 17–69, 24–60, and 1–2.

Techniques:

Journal: PLoS ONE

Article Title: Evaluation of a Novel Hexavalent Humanized Anti-IGF-1R Antibody and Its Bivalent Parental IgG in Diverse Cancer Cell Lines

doi: 10.1371/journal.pone.0044235

Figure Lengend Snippet: IGF-1R expression as determined by flow cytometry.

Article Snippet: Additional anti-human IGF-1R antibodies were obtained from Calbiochem for Ab-1 (clone αIR3), Ab-3 (Clone 33255.111), and Ab-4 (clone 24–31); from Santa Cruz Biotechnology for 2C8, 1H7, 3B7, and C-20; from Millipore for 24–57 and 24–31; and from Invitrogen for 17–69, 24–60, and 1–2.

Techniques: Expressing, Cytometry

Journal: PLoS ONE

Article Title: Evaluation of a Novel Hexavalent Humanized Anti-IGF-1R Antibody and Its Bivalent Parental IgG in Diverse Cancer Cell Lines

doi: 10.1371/journal.pone.0044235

Figure Lengend Snippet: (A) hR1, Hex-hR1, but not hRS7 (irrelevant IgG) induced phosphorylation of IGF-1Rβ and Akt in MCF7 cells in the absence of IGF-1; (B) under the same conditions as in (A), only Hex-hR1 was effective in inducing phosphorylation of IGF-1R1Rβ and Akt in RH-30 cells; (C) MCF7 and RH-30 cells were treated as in (A) except with the addition of IGF-1.

Article Snippet: Additional anti-human IGF-1R antibodies were obtained from Calbiochem for Ab-1 (clone αIR3), Ab-3 (Clone 33255.111), and Ab-4 (clone 24–31); from Santa Cruz Biotechnology for 2C8, 1H7, 3B7, and C-20; from Millipore for 24–57 and 24–31; and from Invitrogen for 17–69, 24–60, and 1–2.

Techniques: Phospho-proteomics

Journal: Cancer science

Article Title: Matrix metalloproteinase-7 triggers the matricrine action of insulin-like growth factor-II via proteinase activity on insulin-like growth factor binding protein 2 in the extracellular matrix.

doi: 10.1111/j.1349-7006.2007.00448.x

Figure Lengend Snippet: Fig. 2. Matrix metalloproteinase (MMP)-7 induced insulin-like growth factor (IGF)-II autocrine response. The IGF type1 receptor (IGF-1R)/Akt signaling pathway was rapidly activated and sustained for at least 180 min by exogenous active MMP-7 in HT29 cells. This IGF-1R phosphorylation was inhibited by TIMP-1(lane7) and IGF-II neutralizing antibody (KM1468, lane8). Cell lysates from HT29 cells (10 µg of protein per lane) were separated by 7.5% (IGF-1R) and 10% (Akt) sodium dodecylsulfate–polyacrylamide gel electrophoresis (SDS-PAGE). The blotted proteins were detected by Western blotting. Bands of phosphorylated IGF-1R (1st antibody, ×500; 2nd antibody, ×6000) were visualized with enhanced chemiluminescence (ECL)-plus chemiluminescent reagent. Bands of total IGF-1R/phosphorylated-Akt/total-Akt (1st antibody, ×500, ×1000, ×1000; 2nd antibody, ×3000) were visualized with ECL reagent. p-, phosphorylated; t-, total.

Article Snippet: The commercially available antibodies used in this study were anti IGFBP-1 mouse monoclonal antibody (Ab) (R & D Systems), anti IGFBP-2 goat polyclonal Ab (C-18, Santa Cruz Biotechnology, Santa Cruz, CA, USA), anti IGFBP3 goat polyclonal Ab (C-19, Santa Cruz Biotechnology), anti IGFBP-4 mouse monoclonal Ab (R & D Systems) anti IGFBP5 goat polyclonal antibody (R & D Systems), anti IGFBP-6 goat polyclonal Ab (C-20, Santa Cruz Biotechnology), anti-IGF-1R rabbit polyclonal Ab (C-20, Santa Cruz Biotechnology), antiphospho-IGF-1R rabbit polyclonal Ab (44–804, BioSource International, Camarillo, CA, USA), anti-Akt rabbit monoclonal Ab (Cell Signaling Technology, Beverly, MA, USA), antiphosphoAkt rabbit monoclonal Ab (Cell Signaling Technology) and peroxidase labeled antimouse/rabbit/goat Abs (Zymed Laboratories, San Francisco, CA, USA).

Techniques: Phospho-proteomics, Polyacrylamide Gel Electrophoresis, SDS Page, Western Blot

Journal: Cancer science

Article Title: Matrix metalloproteinase-7 triggers the matricrine action of insulin-like growth factor-II via proteinase activity on insulin-like growth factor binding protein 2 in the extracellular matrix.

doi: 10.1111/j.1349-7006.2007.00448.x

Figure Lengend Snippet: Fig. 3. Matrix metalloproteinase (MMP)-7 acts as a trigger for the matricrine action of insulin-like growth factor (IGF)-II. (a) The supernatant fraction of extracellular matrix (ECM) from HT29 cells after MMP-7 treatment (200 ng/mL at 37°C for 30 min) was used to determine whether MMP-7 activates IGF-II in the ECM. After residual MMP-7 activity was neutralized by addition of TIMP-1, the supernatant fraction was added to serum-starved HT29 cells. (b) Cell lysates from HT29 cells (10 µg of protein per lane) were separated by 7.5% (IGF-1R) and 10% (Akt) sodium dodecylsulfate–polyacrylamide gel electrophoresis (SDS-PAGE). The blotted proteins were detected by Western blotting. Bands of phosphorylated IGF-1R (1st antibody, ×500; 2nd antibody, ×6000) were visualized with enhanced chemiluminescence (ECL)-plus chemiluminescent reagent. Bands of total IGF-1R/phosphorylated- Akt/total-Akt (1st antibody, ×500, ×1000, ×1000; 2nd antibody, ×3000) were visualized with ECL chemiluminescent reagent. p-, phosphorylated; t-, total; sup, supernatant; SW, SW1116 cells.

Article Snippet: The commercially available antibodies used in this study were anti IGFBP-1 mouse monoclonal antibody (Ab) (R & D Systems), anti IGFBP-2 goat polyclonal Ab (C-18, Santa Cruz Biotechnology, Santa Cruz, CA, USA), anti IGFBP3 goat polyclonal Ab (C-19, Santa Cruz Biotechnology), anti IGFBP-4 mouse monoclonal Ab (R & D Systems) anti IGFBP5 goat polyclonal antibody (R & D Systems), anti IGFBP-6 goat polyclonal Ab (C-20, Santa Cruz Biotechnology), anti-IGF-1R rabbit polyclonal Ab (C-20, Santa Cruz Biotechnology), antiphospho-IGF-1R rabbit polyclonal Ab (44–804, BioSource International, Camarillo, CA, USA), anti-Akt rabbit monoclonal Ab (Cell Signaling Technology, Beverly, MA, USA), antiphosphoAkt rabbit monoclonal Ab (Cell Signaling Technology) and peroxidase labeled antimouse/rabbit/goat Abs (Zymed Laboratories, San Francisco, CA, USA).

Techniques: Activity Assay, Polyacrylamide Gel Electrophoresis, SDS Page, Western Blot

Journal: Cancer science

Article Title: Matrix metalloproteinase-7 triggers the matricrine action of insulin-like growth factor-II via proteinase activity on insulin-like growth factor binding protein 2 in the extracellular matrix.

doi: 10.1111/j.1349-7006.2007.00448.x

Figure Lengend Snippet: Fig. 4. Matrix metalloproteinase (MMP)-7 decreased binding of the insulin-like growth factor (IGF)-II/insulin-like growth factor binding protein (IGFBP)-2 complex to heparan sulfate proteoglycan in the extracellular matrix (ECM) and released bioactive IGF-II. (a) MMP-7 (200 ng/mL) and heparinase III (0.2 U/mL) released IGFBP-2 from the ECM. ECM protein derived from one 6-cm dish was used in each lane. Samples were separated by 15% sodium dodecylsulfate–polyacrylamide gel electrophoresis (SDS-PAGE) and analyzed by Western blotting. Bands of IGFBP-2 (1st antibody, ×500; 2nd antibody, ×3000) were visualized with enhanced chemiluminescence (ECL) reagent. (b) Supernatant from MMP-7-treated (200 ng/mL at 37°C for 30 min) ECM induced phosphorylation of IGF type-1 receptor (IGF-1R), but supernatant from heparinase-III-treated (0.2 U/mL at 37°C for 3 h) ECM did not. 3T3-IGF-1R cells were used in the assay. Cell lysates from HT29 cells (10 µg of protein per lane) were separated by 7.5% SDS- PAGE and analyzed by Western blotting. Bands of phosphorylated IGF-1R (1st antibody, ×500; 2nd antibody, ×6000) were visualized with ECL-plus chemiluminescent reagent. Bands of total IGF-1R (1st antibody, ×500; 2nd antibody, ×3000) were visualized with ECL chemiluminescent reagent. p-, phosphorylated; sup, supernatant; t-, total.

Article Snippet: The commercially available antibodies used in this study were anti IGFBP-1 mouse monoclonal antibody (Ab) (R & D Systems), anti IGFBP-2 goat polyclonal Ab (C-18, Santa Cruz Biotechnology, Santa Cruz, CA, USA), anti IGFBP3 goat polyclonal Ab (C-19, Santa Cruz Biotechnology), anti IGFBP-4 mouse monoclonal Ab (R & D Systems) anti IGFBP5 goat polyclonal antibody (R & D Systems), anti IGFBP-6 goat polyclonal Ab (C-20, Santa Cruz Biotechnology), anti-IGF-1R rabbit polyclonal Ab (C-20, Santa Cruz Biotechnology), antiphospho-IGF-1R rabbit polyclonal Ab (44–804, BioSource International, Camarillo, CA, USA), anti-Akt rabbit monoclonal Ab (Cell Signaling Technology, Beverly, MA, USA), antiphosphoAkt rabbit monoclonal Ab (Cell Signaling Technology) and peroxidase labeled antimouse/rabbit/goat Abs (Zymed Laboratories, San Francisco, CA, USA).

Techniques: Binding Assay, Derivative Assay, Polyacrylamide Gel Electrophoresis, SDS Page, Western Blot, Phospho-proteomics

Journal: Cancer science

Article Title: Matrix metalloproteinase-7 triggers the matricrine action of insulin-like growth factor-II via proteinase activity on insulin-like growth factor binding protein 2 in the extracellular matrix.

doi: 10.1111/j.1349-7006.2007.00448.x

Figure Lengend Snippet: Fig. 7. Recovery of insulin-like growth factor (IGF)-II mediated phosphorylation of IGF type1 receptor (IGF-1R) through Insulin-like growth factor binding protein (IGFBP)-2 degradation by matrix metalloproteinase (MMP)-7. Cell lysates from HT29 cells (10 µg of protein per lane) exposed to IGF system component molecules (IGF-II/IGFBP-2/MMP-7, preincubation conditions; see ‘Materials and Methods’) were separated by 7.5% sodium dodecylsulfate–polyacrylamide gel electrophoresis (SDS-PAGE) and analyzed by Western blotting. Bands of phosphorylated IGF-1R (1st antibody, ×500; 2nd antibody, ×6000) were visualized with enhanced chemiluminescence (ECL)-plus chemiluminescent reagent. Bands of total IGF-1R (1st antibody, ×500; 2nd antibody, ×3000) were visualized with ECL chemiluminescent reagent. p-, phosphorylated; t-, total.

Article Snippet: The commercially available antibodies used in this study were anti IGFBP-1 mouse monoclonal antibody (Ab) (R & D Systems), anti IGFBP-2 goat polyclonal Ab (C-18, Santa Cruz Biotechnology, Santa Cruz, CA, USA), anti IGFBP3 goat polyclonal Ab (C-19, Santa Cruz Biotechnology), anti IGFBP-4 mouse monoclonal Ab (R & D Systems) anti IGFBP5 goat polyclonal antibody (R & D Systems), anti IGFBP-6 goat polyclonal Ab (C-20, Santa Cruz Biotechnology), anti-IGF-1R rabbit polyclonal Ab (C-20, Santa Cruz Biotechnology), antiphospho-IGF-1R rabbit polyclonal Ab (44–804, BioSource International, Camarillo, CA, USA), anti-Akt rabbit monoclonal Ab (Cell Signaling Technology, Beverly, MA, USA), antiphosphoAkt rabbit monoclonal Ab (Cell Signaling Technology) and peroxidase labeled antimouse/rabbit/goat Abs (Zymed Laboratories, San Francisco, CA, USA).

Techniques: Phospho-proteomics, Binding Assay, Polyacrylamide Gel Electrophoresis, SDS Page, Western Blot