Whole hippocampal tissues from P9 mice were homogenized using a glass homogenizer (Wheaton) in ice cold homogenization buffer (10 mM Tris-HCl, pH7.4, 1% SDS, 1% protease inhibitor cocktail (#8340, Sigma)), 1% phosphatase inhibitor 2 (#5726, Sigma). The homogenate was centrifuged for 15 min at 1000 × g at 4 °C, and the supernatant diluted with 5x final sample buffer and centrifuged at 13,000 g. Forty microgram total protein was loaded per lane on a 7.5% acrylamide/bis gel, and transferred to nitrocellulose membrane. After blocking with blotto (5% #9999S Cell Signaling in phosphate-buffered saline), anti-MET antibody (#8057, 1:3000, Santa Cruz Biotechnology), and secondary antibody (#715-035-150, 1:5000, Jackson ImmunoResearch) was used for immunodetection, followed by Femto chemiluminescent substrate (#34095, ThermoFisher). The signal was analyzed using a CCD camera (UVP BioImaging System) and VisionWorksLS software (VisionWorks). The immunostaining process was repeated for anti-α-Tubulin protein (#CP06, 1:200,000, EMD Millipore) to normalize anti-MET signal.
Anti met antibody
Manufactured by Santa Cruz Biotechnology
The Anti-MET antibody is a research tool used to detect and study the MET protein, also known as hepatocyte growth factor receptor. It is a highly specific and sensitive antibody that can be utilized in various experimental techniques such as Western blotting, immunoprecipitation, and immunohistochemistry to analyze the expression and localization of the MET protein.
Automatically generated - may contain errors
2 protocols using anti met antibody
1
Quantifying Hippocampal MET Protein
2
Analyzing Receptor Tyrosine Kinase Activation
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Cells were seeded on 100-mm-diameter fibronectin-coated tissue culture dishes, grown to semi-confluency, and treated either with HGF or HGF and cloudberry extract. Cell lysates were prepared as described above except that cells were lysed with 1000 μL of lysis buffer per sample, containing 10% of glycerol. 250 μg of protein from the lysates were incubated with 25 μL of anti-Met antibody conjugated to protein A/G PLUS-agarose (Santa Cruz) for 4 h at 4°C. The pellets were collected by centrifugation at 1000 × g and washed four times for 5 min each with lysis buffer at 4°C. The pellets were resuspended in 30 μL of Laemmli sample buffer, heated at 98°C for 5 min, resolved on 7.5% SDS-PAGE and transferred to PVDF membrane. The blots were probed with anti-phosphotyrosine antibody (Clone 4G10, Upstate), followed by stripping and probing for total Met levels with anti-Met antibody (Santa Cruz). The blots were developed and bands quantified as described above.
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