Cells were harvested and lysed in NETN (20 mM Tris-HCl, pH 8.0, 100 mM NaCl, 1 mM EDTA, 0.5% NP40) for 10 min on ice. Lysates were cleared by centrifugation at 13,200 rpm at 4°C for 10 min. Supernatants were collected and protein concentrations were determined by the Bradford assay (Bio-Rad). The proteins were then separated with a SDS/polyacrylamide gel and transferred to a Nitrocellulose membrane (Bio-Rad). After blocking in TBS with 5% BSA (Sigma) for 1 hr, the membranes were incubated overnight at 4-8°C with the primary antibodies in TBS containing 1% BSA. The following antibodies were utilized: the β-actin antibody was purchased from Sigma; and the tubulin, B7H3, B-Raf, MEK and MVP antibody were purchased from Santa Cruz; MEK-P and Phospho-Tyrosine was purchased from cell signaling. Membranes were extensively washed with TBST and incubated with horseradish peroxidase conjugated secondary anti-mouse antibody or anti-rabbit antibody (dilution 1:2,500, Bio-Rad). After additional washes with TBST, antigen-antibody complexes were visualized with the enhanced chemiluminescence kit (Pierce).
Mek p and phospho tyrosine
Manufactured by Cell Signaling Technology
MEK-P is an antibody that detects phosphorylated MEK1/2, a key component of the MAPK signaling pathway. Phospho-Tyrosine is an antibody that detects phosphorylated tyrosine residues on proteins. These products are intended for use in Western blotting, immunohistochemistry, and other immunoassay applications to study cell signaling processes.
Automatically generated - may contain errors
Lab products found in correlation
Horseradish peroxidase conjugated secondary anti mouse antibody or anti rabbit antibody, by Bio-Rad (2 mentions)
β actin antibody, by Merck Group (2 mentions)
Bovine serum albumin (bsa), by Merck Group (2 mentions)
Nitrocellulose membrane, by Bio-Rad (2 mentions)
Bradford assay, by Bio-Rad (2 mentions)
2 protocols using mek p and phospho tyrosine
1
Western Blot Analysis of Cell Signaling
2
Western Blot Analysis of Cell Signaling
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Cells were harvested and lysed in NETN (20 mM Tris-HCl, pH 8.0, 100 mM NaCl, 1 mM EDTA, 0.5% NP40) for 10 min on ice. Lysates were cleared by centrifugation at 13,200 rpm at 4°C for 10 min. Supernatants were collected and protein concentrations were determined by the Bradford assay (Bio-Rad). The proteins were then separated with a SDS/polyacrylamide gel and transferred to a Nitrocellulose membrane (Bio-Rad). After blocking in TBS with 5% BSA (Sigma) for 1 hr, the membranes were incubated overnight at 4-8°C with the primary antibodies in TBS containing 1% BSA. The following antibodies were utilized: the β-actin antibody was purchased from Sigma; and the tubulin, B7H3, B-Raf, MEK and MVP antibody were purchased from Santa Cruz; MEK-P and Phospho-Tyrosine was purchased from cell signaling. Membranes were extensively washed with TBST and incubated with horseradish peroxidase conjugated secondary anti-mouse antibody or anti-rabbit antibody (dilution 1:2,500, Bio-Rad). After additional washes with TBST, antigen-antibody complexes were visualized with the enhanced chemiluminescence kit (Pierce).
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