Anti-IGF1 Receptor antibody [EPR19322]

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-IGF1 Receptor antibody [EPR19322] (AB182408)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol permeabilized C2C12 (Mouse myoblast cell line) cells labeling IGF1 Receptor with ab182408 at 1/80 dilution (red) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat anti Rabbit IgG (Alexa Fluor^® 488) at 1/500 dilution was used as the secondary antibody.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-IGF1 Receptor antibody [EPR19322] (AB182408)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized C2C12 (Mouse myoblast cell line) cells labeling IGF1 Receptor with ab182408 at 1/1000 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing membranous and cytoplasmic staining on C2C12 cell line. The nuclear counter stain is DAPI (blue).

Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed (ab150120) at 1/1000 dilution (red).

The negative controls are as follows :

-ve control 1 : ab182408 at 1/1000 dilution followed by ab150120 at 1/1000 dilution.
-ve control 2 : ab7291 at 1/1000 dilution followed by ab150077 at 1/1000 dilution.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-IGF1 Receptor antibody [EPR19322] (AB182408)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized C6 (Rat glial tumor cell line) cells labeling IGF1 Receptor with ab182408 at 1/1000 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing membranous and cytoplasmic staining on C6 cell line. The nuclear counter stain is DAPI (blue).

Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed (ab150120) at 1/1000 dilution (red).

The negative controls are as follows :

-ve control 1 : ab182408 at 1/1000 dilution followed by ab150120 at 1/1000 dilution.
-ve control 2 : ab7291 at 1/1000 dilution followed by ab150077 at 1/1000 dilution.

• IP

Supplier Data

Immunoprecipitation - Anti-IGF1 Receptor antibody [EPR19322] (AB182408)

IGF1 Receptor was immunoprecipitated from 1mg of C2C12 (Mouse myoblast cell line) whole cell lysate with ab182408 at 1/50 dilution. Western blot was performed from the immunoprecipitate using ab182408 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

Lane 1 : C2C12 whole cell lysate, 10μg (Input).

Lane 2 : ab182408 IP in C2C12 whole cell lysate.

Lane 3 : Rabbit IgG,monoclonal [EPR25A] - Isotype Control (ab172730) instead of ab182408 in C2C12 whole cell lysate.

Blocking and dilution buffer and concentration : 5% NFDM/TBST.

Exposure time : 3 seconds.

All lanes:

Immunoprecipitation - Anti-IGF1 Receptor antibody [EPR19322] (ab182408)

Predicted band size: 154 kDa

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• WB

Supplier Data

Western blot - Anti-IGF1 Receptor antibody [EPR19322] (AB182408)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

This blot was developed using a high sensitivity ECL substrate. The high-sensitivity ECL substrate used allows for the detection of proteins in the mid-femtogram range.

In Western blot, Anti-IGF1 Receptor antibody [EPR19322] (ab182408) staining at 1/1000 dilution

All lanes:

Western blot - Anti-IGF1R (pY1161) + INSR (pY1185) + INSRR (pY1141) antibody [EPR26123-29] (<a href='/en-us/products/primary-antibodies/igf1r-py1161-insr-py1185-insrr-py1141-antibody-epr26123-29-ab314116'>ab314116</a>) at 1/1000 dilution

Lane 1:

HeLa (human cervix adenocarcinoma epithelial cell) starved for 18 hours whole cell lysate (untreated membrane) at 30 µg

Lane 2:

HeLa starved for 18 hours, then treated with 100ng/mL IGF-1 for 10 minutes whole cell lysate (untreated membrane) at 30 µg

Lane 3:

HeLa starved for 18 hours whole cell lysate (alkaline phosphatase treated membrane) at 30 µg

Lane 4:

HeLa starved for 18 hours, then treated with 100ng/mL IGF-1 for 10 minutes whole cell lysate (alkaline phosphatase treated membrane) at 30 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 100 kDa

true

Exposure time: 81s

• WB

Lab

Western blot - Anti-IGF1 Receptor antibody [EPR19322] (AB182408)

False colour image of Western blot : Anti-IGF1 Receptor antibody [EPR19322] staining at 1/1000 dilution, shown in green; Mouse anti-CANX [CANX/1543] (ab238078) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab182408 was shown to bind specifically to IGF1 Receptor. A band was observed at 82 kDa (beta chain) in wild-type MCF7 cell lysates with no signal observed at this size in IGF1R knockout cell line. To generate this image, wild-type and IGF1R knockout MCF7 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with primary antibodies overnight at 4°C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-IGF1 Receptor antibody [EPR19322] (ab182408) at 1/1000 dilution

Lane 1:

Wild-type MCF7 cell lysate at 20 µg

Lane 2:

IGF1R knockout MCF7 cell lysate at 20 µg

Lane 3:

HeLa cell lysate at 20 µg

Lane 4:

HDLM-2 cell lysate at 20 µg

Observed band size: 82 kDa

false

• WB

Lab

Western blot - Anti-IGF1 Receptor antibody [EPR19322] (AB182408)

Lane 1 : Wild-type HAP1 whole cell lysate (40 μg)
Lane 2 : IGF1R knockout HAP1 whole cell lysate (40 μg)
Lane 3 : HeLa whole cell lysate (40 μg)

Lanes 1 - 3 Merged signal (red and green). Green - ab182408 observed at 100 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab182408 was shown to specifically recognize IGF1R in wild-type HAP1 cells along with additional cross-reactve bands. No band was observed when IGF1R knockout samples were examined. Wild-type and IGF1R knockout samples were subjected to SDS-PAGE. ab182408 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1/2000 dilution and 1/10,000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed ab216776 secondary antibodies at 1/10,000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-IGF1 Receptor antibody [EPR19322] (ab182408)

Predicted band size: 154 kDa

false

• WB

Supplier Data

Western blot - Anti-IGF1 Receptor antibody [EPR19322] (AB182408)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

This blot was developed using a high sensitivity ECL substrate. The high-sensitivity ECL substrate used allows for the detection of proteins in the mid-femtogram range.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

In Western blot, Anti-IGF1 Receptor antibody [EPR19322] (ab182408) staining at 1/1000 dilution.

All lanes:

Western blot - Anti-IGF1R (pY1161) + INSR (pY1185) + INSRR (pY1141) antibody [EPR26123-29] (<a href='/en-us/products/primary-antibodies/igf1r-py1161-insr-py1185-insrr-py1141-antibody-epr26123-29-ab314116'>ab314116</a>) at 1/1000 dilution

Lane 1:

HeLa (human cervix adenocarcinoma epithelial cell) starved for 18 hours whole cell lysate at 30 µg

Lane 2:

HeLa starved for 18 hours, then treated with 100ng/ml IGF-1 for 5 minutes whole cell lysate at 30 µg

Lane 3:

HeLa starved for 18 hours, then treated with 100ng/ml IGF-1 for 20 minutes whole cell lysate at 30 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 100 kDa

true

Exposure time: 92s

• WB

Lab

Western blot - Anti-IGF1 Receptor antibody [EPR19322] (AB182408)

Anti-IGF1R antibody [EPR19322] (ab182408) staining at 1/1000 dilution, shown in green; Mouse anti-CANX [CANX/1543] (ab238078) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab182408 was shown to bind specifically to IGF1R. A band was observed at 82 kDa in wild-type HCT 116 cell lysates with no signal observed at this size in IGF1R knockout cell line. To generate this image, wild-type and IGF1R knockout HCT 116 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-IGF1 Receptor antibody [EPR19322] (ab182408) at 1/1000 dilution

Lanes 1 - 4:

Western blot at 20 µg

Lane 2:

Western blot - Human IGF1R knockout HCT116 cell line (<a href='/en-us/products/cell-lines/human-igf1r-knockout-hct116-cell-line-ab287509'>ab287509</a>)

Secondary

All lanes:

Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

Predicted band size: 154 kDa

Observed band size: 82 kDa

false

• WB

Lab

Western blot - Anti-IGF1 Receptor antibody [EPR19322] (AB182408)

Lanes 1- 2 : Merged signal (red and green). Green - ab182408 observed at 100 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.

ab182408 was shown to react with IGF1 Receptor in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab264801 (knockout cell lysate ab256951) was used. Wild-type HeLa and IGF1R knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab182408 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^®800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^®680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-IGF1 Receptor antibody [EPR19322] (ab182408) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

IGF1R knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human IGF1R (IGF1 Receptor) knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-igf1r-igf1-receptor-knockout-hela-cell-line-ab264801'>ab264801</a>)

Predicted band size: 154 kDa

Observed band size: 100 kDa

false

• WB

Supplier Data

Western blot - Anti-IGF1 Receptor antibody [EPR19322] (AB182408)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : Lane 1 : 5 seconds; Lane 2-4 : 3 minutes; Lane 5/6 : 2 seconds.

The expression profile observed is consistent with what has been described in the literature (PMID : 11402025).

Lanes 1 - 4:

Western blot - Anti-IGF1 Receptor antibody [EPR19322] (ab182408) at 1/2000 dilution

Lanes 5 - 6:

Western blot - Anti-IGF1 Receptor antibody [EPR19322] (ab182408) at 1/1000 dilution

Lane 1:

C2C12 (Mouse myoblast cell line) whole cell lysate at 20 µg

Lane 2:

HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg

Lane 3:

293 (Human epithelial cell line from embryonic kidney) whole cell lysate at 20 µg

Lane 4:

MCF7 (Human breast adenocarcinoma cell line) whole cell lysate at 20 µg

Lane 5:

C6 (Rat glial tumor cell line) whole cell lysate at 10 µg

Lane 6:

NIH/3T3 (Mouse embryonic fibroblast cell line) whole cell lysate at 10 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 154 kDa

Observed band size: 100 kDa,200 kDa

false

• WB

Supplier Data

Western blot - Anti-IGF1 Receptor antibody [EPR19322] (AB182408)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : Lane 1 : 10 seconds; Lane 2 : 3 minutes; Lane 3 : 1 minute.

The expression profile observed is consistent with what has been described in the literature (PMID : 11402025).

Lane 1:

Western blot - Anti-IGF1 Receptor antibody [EPR19322] (ab182408) at 1/1000 dilution

Lanes 2 - 3:

Western blot - Anti-IGF1 Receptor antibody [EPR19322] (ab182408) at 1/5000 dilution

Lane 1:

Mouse brain lysate at 10 µg

Lane 2:

Mouse spleen lysate at 10 µg

Lane 3:

Rat brain lysate at 10 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 154 kDa

Observed band size: 100 kDa,200 kDa

false

• WB

CiteAb

Western blot - Anti-IGF1 Receptor antibody [EPR19322] (AB182408)

IGF1 Receptor western blot using anti-IGF1 Receptor antibody [EPR19322] ab182408. Publication image and figure legend from Su, M., Fan, S., et al., 2020, Front Physiol, PubMed 32733269.

ab182408 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab182408 please see the product overview.

MiR-223 directly targets the IGF-1R, which inhibits the phosphorylation of Akt and GSK3β. (A) Bioinformatics analysis identified a binding site for miR-223 in the 3'-UTR of IGF-1R. Luciferase activity assay in VSMCs following introduction of IGF-1R 3' UTR or mutant 3' UTR (mut) with or without miR-223 mimic (n = 3). VSMCs were transfected with miR-223 mimic for 48 h. (B,C) The expression of IGF-1R in VSMC was detected with western blot analysis. (D) The rate of proliferation was determined by CCK8 assay in VSMCs transfected with overexpression of IGF-1R. (E,F) The phosphorylation of Akt and GSK3β in VSMC was determined with western blot analysis. Data are presented as mean ± SD (n = 6). Comparisons between two groups were analyzed using unpaired nonparametric Student's t-test. *p < 0.05, **p < 0.01, ***p < 0.001 vs Ctrl (miR-NC).

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