Anti-Met (c-Met) antibody [EP1454Y] - N-terminal

• WB

Lab

Western blot - Anti-Met (c-Met) antibody [EP1454Y] - N-terminal (AB51067)

False colour image of Western blot : Anti-Met (c-Met) antibody [EP1454Y] - N-terminal staining at 1/1000 dilution, shown in green; Rabbit Anti-GAPDH antibody [EPR16891] (ab181602) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab51067 was shown to bind specifically to the alpha chain of c-Met. A band was observed at 50 kDa in wild-type HeLa cell lysates with no signal observed at this size in MET knockout cell line ab265961 (knockout cell lysate ab256991). To generate this image, wild-type and MET knockout HeLa cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween^® 20 (TBS-T) 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 HRP conjugated Goat anti-Rabbit (H+L) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

All lanes:

Western blot - Anti-Met (c-Met) antibody [EP1454Y] - N-terminal (ab51067) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

MET knockout HeLa cell lysate at 20 µg

Predicted band size: 155 kDa

Observed band size: 50 kDa

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• WB

Lab

Western blot - Anti-Met (c-Met) antibody [EP1454Y] - N-terminal (AB51067)

Western blot : Anti-MET antibody [EP1454Y] (ab51067) staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab51067 was shown to bind specifically to MET. A band was observed at 40-50 kDa in wild-type A549 cell lysates with no signal observed at this size in MET knockout cell line. To generate this image, wild-type and MET knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3% milk in TBS-0.1% Tween® 20 (TBS-T) 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.

Lanes 1 - 6:

Western blot - Anti-Met (c-Met) antibody [EP1454Y] - N-terminal (ab51067) at 1/1000 dilution

Lanes 1 - 2:

Western blot - Anti-IMPDH2 antibody [EPR8365(B)] (<a href='/en-us/products/primary-antibodies/impdh2-antibody-epr8365b-ab129165'>ab129165</a>) at 1/1000 dilution

Lane 1:

Wild-type A549 cell lysate at 20 µg

Lane 2:

MET knockout A549 cell lysate at 20 µg

Lane 3:

Wild-type HeLa ab255929 cell lysate at 20 µg

Lane 4:

MET (Met (c-Met)) knockout HeLa <a href='/en-us/products/cell-lysates/human-met-c-met-knockout-hela-cell-lysate-ab256991'>ab256991</a> cell lysate at 20 µg

Lane 5:

HT-29 cell lysate at 20 µg

Lane 6:

T-47D cell lysate at 20 µg

Secondary

All lanes:

NFDM/TBST at 1/20000 dilution

Observed band size: 40-50 kDa

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• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Met (c-Met) antibody [EP1454Y] - N-terminal (AB51067)

Immunohistochemical staining of paraffin embedded human bladder carcinoma with purified ab51067 at a working dilution of 1/100. The secondary antibody used is HRP goat anti-rabbit IgG H&L (ab97051) at 1/500. The sample is counterstained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0.

PBS was used instead of the primary antibody as the negative control (inset).

• IHC-P

AbReview60220****

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Met (c-Met) antibody [EP1454Y] - N-terminal (AB51067)

ab51067 staining Met (c-Met) in human breast tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections).

Tissue was fixed with formaldehyde, permeabilized with 0.05% Tween-20 and blocked for 30 minutes at 22°C; antigen retrieval was by heat mediation in antigen retrieval buffer (100X citrate buffer pH 6.0) (ab94674). Samples were incubated with the primary antibody (1/100) for 14 hours at 4°C. An Alexa Fluor^® 488-conjugated goat anti-rabbit IgG polyclonal (1/300) was used as the secondary antibody.

This image is courtesy of an Abreview submitted by David Ivancic.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Met (c-Met) antibody [EP1454Y] - N-terminal (AB51067)

Immunohistochemical staining of paraffin embedded human clear cell kidney carcinoma with purified ab51067 at a working dilution of 1/100. The secondary antibody used is HRP goat anti-rabbit IgG H&L (ab97051) at 1/500. The sample is counterstained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0.

PBS was used instead of the primary antibody as the negative control (inset).

• I-ELISA

Supplier Data

Indirect ELISA - Anti-Met (c-Met) antibody [EP1454Y] - N-terminal (AB51067)

ELISA analysis of Human Met recombinant protein at 1000 ng/mL with ab51067. An Alkaline Phosphatase-conjugated AffiniPure Goat Anti-Rabbit IgG (H+L) at 1/2500 dilution was used as the secondary antibody.

• WB

Lab

Western blot - Anti-Met (c-Met) antibody [EP1454Y] - N-terminal (AB51067)

Lanes 1 - 3 : Merged signal (red and green). Green - ab51067 observed at 50 kDa. Red - loading control, ab18058, observed at 130 kDa.

This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using Licor blocking buffer before being incubated with ab51067 and ab18058 (loading control) overnight at 4°C. Antibody binding was detected using Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at a 1 : 10000 dilution for 1hr at room temperature and then imaged.

All lanes:

Western blot - Anti-Met (c-Met) antibody [EP1454Y] - N-terminal (ab51067) at 1/1000 dilution

Lane 1:

Mouse thymus tissue lysate at 20 µg

Lane 2:

Rat thymus tissue lysate at 20 µg

Lane 3:

Mouse lung tissue lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution

Predicted band size: 155 kDa

Observed band size: 50 kDa

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• WB

Supplier Data

Western blot - Anti-Met (c-Met) antibody [EP1454Y] - N-terminal (AB51067)

Lanes 1 - 2 : Merged signal (red and green). Green - ab216574 observed at 156 kDa. Red - loading control, ab8245 observed at 37 kDa.

ab216574 was shown to react with Met (c-Met) in wild-type HeLa. Loss of signal was observed when knockout cell line ab265961 (knockout cell lysate ab256991) was used. Wild-type and Met (c-Met) knockout samples were subjected to SDS-PAGE. ab216574 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 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-Met (c-Met) antibody [EP1454Y] - N-terminal (ab51067) at 1/1000 dilution

Lane 1:

Wild-type HeLa at 20 µg

Lane 2:

MET knockout HeLa at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Predicted band size: 155 kDa

Observed band size: 37 kDa

false

• WB

Lab

Western blot - Anti-Met (c-Met) antibody [EP1454Y] - N-terminal (AB51067)

Blocking buffer / Diluent and concentration :

5% NFDM/TBST

All lanes:

Western blot - Anti-Met (c-Met) antibody [EP1454Y] - N-terminal (ab51067) at 1/1000 dilution

All lanes:

HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates at 20 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 1/10000 dilution

Predicted band size: 155 kDa

Observed band size: 50 kDa

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Exposure time: 60s

• WB

CiteAb

Western blot - Anti-Met (c-Met) antibody [EP1454Y] - N-terminal (AB51067)

Met (c-Met) western blot using anti-Met (c-Met) antibody [EP1454Y] - N-terminal ab51067. Publication image and figure legend from Zhao, Q., Zhu, H. P., et al., 2020, Int J Mol Sci, PubMed 32156008.

ab51067 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 ab51067 please see the product overview.

(A) Cells were incubated with various concentrations (0, 20, or 40 μg/mL) of DHP1808 for 24 h. Expression or phosphorylation levels of HSP90 client proteins were determined by western blot analysis; (B) Cells were incubated with various concentrations (0, 20, or 40 μg/mL) of DHP1808 for 24 h. Expression or phosphorylation levels of proteins associated with MAPK signaling pathway were determined by western blot analysis; (C) Cells were incubated with various concentrations (0, 20, or 40 μg/mL) of DHP1808 for 24 h. Expression or phosphorylation level of proteins associated with the PI3K-Akt signaling pathway was determined by western blot analysis; (D). The fold change of EGFR expression was determined by qPCR analysis.

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