MA5-13319

antibody from Invitrogen Antibodies

Targeting: EGFR ERBB, ERBB1

Provider product page for MA5-13319

Western blot

Immunocytochemistry

Flow cytometry

Other assay

Antibody data

Antibody Data
Antigen structure
References [64]
Comments [0]
Validations
Western blot [1]
Immunocytochemistry [5]
Flow cytometry [1]
Other assay [6]

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Product number: MA5-13319 - Provider product page
Provider: Invitrogen Antibodies
Product name: EGFR Monoclonal Antibody (199.12)
Antibody type: Monoclonal
Antigen: Recombinant full-length protein
Description: MA5-13319 targets Epidermal Growth Factor Receptor in IP, ICC/IF and WB applications and shows reactivity with Human samples.
Antibody clone number: 199.12
Concentration: 0.2 mg/mL

EGFR protein structure - MA5-13319 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of EGFR (green) showing staining in the cytoplasm and membrane of A431 cells. Formalin-fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 5-10 minutes and blocked with 3% BSA-PBS for 30 minutes at room temperature. Cells were probed with a Epidermal Growth Factor Receptor monoclonal antibody (Product # MA5-13319) in 3% BSA-PBS at a dilution of 1:50 and incubated overnight at 4 ºC in a humidified chamber. Cells were washed with PBST and incubated with a DyLight-conjugated secondary antibody in PBS at room temperature in the dark. F-actin (red) was stained with a fluorescent red phalloidin and nuclei (blue) were stained with Hoechst or DAPI. Images were taken at a magnification of 60x.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of EGFR (green) showing staining in the cytoplasm and membrane of HeLa cells. Formalin-fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 5-10 minutes and blocked with 3% BSA-PBS for 30 minutes at room temperature. Cells were probed with a Epidermal Growth Factor Receptor monoclonal antibody (Product # MA5-13319) in 3% BSA-PBS at a dilution of 1:50 and incubated overnight at 4 ºC in a humidified chamber. Cells were washed with PBST and incubated with a DyLight-conjugated secondary antibody in PBS at room temperature in the dark. F-actin (red) was stained with a fluorescent red phalloidin and nuclei (blue) were stained with Hoechst or DAPI. Images were taken at a magnification of 60x.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of EGFR was performed using 90% confluent log phase A-431 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with EGFR Mouse monoclonal antibody (Product # MA5-13319) at 5 µg/mL in 0.1% BSA and incubated for 3 hours at room temperature and then labeled with Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing membranous localization. Panel e shows the no primary antibody control. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of EGFR was done on 90% confluent log phase A431 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled EGFR (199.12) Mouse Monoclonal Antibody (Product # MA5-13319) at 2 µg/mL in 0.1% BSA and incubated for 3 hours at room temperature and then labeled with Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Alexa Fluor® 555 Rhodamine Phalloidin (Product # R415, 1:300). Panel d is a merged image showing membranous localization. Panel e is a no primary antibody control. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of EGFR was performed using 70% confluent log phase A-431 cells (WIld type, panels a,d), CAS9 control (panels b,e) and EGFR Knockout (panels c,f). The cells were fixed, permeabilized, and labelled with EGFR Mouse Monoclonal Antibody(Product # MA5-13319, 5 µg/mL), followed by Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175, 1:2000). Nuclei (blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938) and Rhodamine Phalloidin (Product # R415, 1:300) was used for cytoskeletal F-actin (red) staining. Loss of signal was observed in EGFR Knockout cells (panel c,f) confirming specificity of the antibody to EGFR(green). The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunoprecipitation of Epidermal Growth Factor Receptor using Epidermal Growth Factor Receptor Monoclonal Antibody (Product # MA5-13319) on Native Human A431 Cells.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 8 Investigation of EGFR and pEGFR. ( A ) Cells were stained for EGFR (red), pEGFR (green), and nuclei (blue) (scale bar: 20 mum). ( B ) Representative images of EGFR clusters with corresponding heat maps of relative intensities. ( C ) (+) EGF demonstrated significantly lower EGFR to cell area ratio. ( D , E ) (+) EGF displayed significantly higher pEGFR to cell area ratio and pEGFR to EGFR area ratio ( p < 0.05 calculated from Student's t -test with n > 18 from more than three devices for each condition).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 1 Characterization of MCF7-derived MCF7-EGFR cells. ( A ) Flow cytometry analysis of surface EGFR expression. Cells from gate P1 were sorted and cultivated as MCF7-EGFR cells. ( B ) Western blot of total cellular EGFR (typical image). Tubulin was used as a loading control.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 4 SCAMP3 colocalizes with EGFR after receptor internalization. Cells were stimulated with 10 ng/mL EGF at the indicated time points to evaluate the location of SCAMP3 and EGFR using confocal microscopy. Representative images of the internalization assay and fluorescence quantification of SCAMP3, EGFR, and their colocalization in ( A - C ) SUM-149 and ( D - F ) MDA-MB-468 cells. The nuclei were stained with DAPI (blue). Secondary antibodies conjugated to Alexa 488 (green) and Alexa 594 (red) were used to detect SCAMP3 and EGFR, respectively. The micrographs were taken at a magnification of 60x using confocal microscopy. The white arrows indicate the colocalization of EGFR and SCAMP3 in the zoom images. The zoom-in of each image is shown in white squares and each has equal dimensions. Scale bar = 20 um. Total cell fluorescence and colocalization area analyses were performed in 20 cells from three experiments using Image J. Colocalization was calculated as the ratio of the colocalization fluorescence area to the total cell fluorescence area. One way or two-way ANOVA; * p

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Characterization of gallbladder tumor cell lines. A Western blot analysis of EGFR expression in NOZ cells (right lane) and HEK293 cells (left lane). B Flow cytometric analysis of fluorescent anti-EGFR bounding in NOZ cells (blue line) and HEK293 cells (red line). C Labeling of NOZ and HEK293 cells with fluorescent anti-EGFR (x40 water immersion objective with the LSM 700 confocal microscope, using 405- and 488-nm lasers). The white scale bar represents 10 mum

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 1--figure supplement 1. Representative immunofluorescence stains of Akt phosphorylation and cell surface EGF receptor levels. Representative immunofluorescence images of MCF10A cells used for the generation of quantitative data on Akt phosphorylation (pAkt) and cell surface EGFR (sEGFR). (left) MCF10A cells 5 min after control treatment ( a ), or treatment with 1 ng/ml ( b ) or 100 ng/ml of EGF stained for pAKT. (right) MCF 10A cells 180 min after control treatment (a), or treatment with 1 ng/ml (b) or 100 ng/ml of EGF stained for sEGFR. Pseudocolors: blue: nuclei; green: cytoplasm; red: pAKT (left panels) or sEGFR (right panels).