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Phospho-EGFR pTyr1086 Antibody (701736) in WB
A) Western blot analysis was performed on whole cell extracts (50 ug lysate) of A431 (Lane 1), A431 treated with EGF (200ng/mL/ 10 minutes)(Lane 2), HeLa (Lane 3), HeLa treated with EGF (200ng/mL/10 minutes) (Lane 4). The blots were probed with ABfinity? EGFR (pY1086) Recombinant Rabbit Monoclonal Antibody (Product# 701736 2-3 ug/ml) and detected by chemiluminescence using Goat Anti-Rabbit IgG (H+L) Superclonal? Secondary Antibody, HRP conjugate (Product # A27036, 0.4 ug/ml, 1:2500 dilution). A 135 kDa band corresponding to EGFR(pY1086) was observed across cell lines according to the treatment. B) To confirm the specificity of ABfinity? Anti-EGFR[pY1086] Recombinant Rabbit Monoclonal Antibody, competition was performed with the phosphopeptide (10 ug/mL) as shown in the corresponding blot on the right. The peptide competes with the antibody and prevents it from binding to the target protein. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 4-12% Bis-Tris gel (Product # NP0321BOX), XCell SureLock? Electrophoresis System (Product # EI0002), and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800).Resolved proteins were then transferred onto a nitrocellulose membrane by overnight wet transfer The membrane was probed with the relevant primary and secondary antibody following blocking with 5% skimmed milk. Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).
![A) Western blot analysis was performed on whole cell extracts (50 ug lysate) of A431 (Lane 1), A431 treated with EGF (200ng/mL/ 10 minutes)(Lane 2), HeLa (Lane 3), HeLa treated with EGF (200ng/mL/10 minutes) (Lane 4). The blots were probed with ABfinity? EGFR (pY1086) Recombinant Rabbit Monoclonal Antibody (Product# 701736 2-3 ug/ml) and detected by chemiluminescence using Goat Anti-Rabbit IgG (H+L) Superclonal? Secondary Antibody, HRP conjugate (Product # A27036, 0.4 ug/ml, 1:2500 dilution). A 135 kDa band corresponding to EGFR(pY1086) was observed across cell lines according to the treatment. B) To confirm the specificity of ABfinity? Anti-EGFR[pY1086] Recombinant Rabbit Monoclonal Antibody, competition was performed with the phosphopeptide (10 ug/mL) as shown in the corresponding blot on the right. The peptide competes with the antibody and prevents it from binding to the target protein. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 4-12% Bis-Tris gel (Product # NP0321BOX), XCell SureLock? Electrophoresis System (Product # EI0002), and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800).Resolved proteins were then transferred onto a nitrocellulose membrane by overnight wet transfer The membrane was probed with the relevant primary and secondary antibody following blocking with 5% skimmed milk. Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).](http://tools.thermofisher.com/content/sfs/gallery/low/701736-EGFRpY1086-WB.jpg)
