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HT 29 cell lysate was prepared by homogenization in modified RIPA buffer (150 mM sodium chloride, 50 mM Tris-HCl, pH 7.4, 1 mM ethylene diamine tetraacetic acid, 1 mM phenylmethylsulfonyl flouride, 1% Triton X-100, 1% sodium deoxycholic acid, 0.1% sodium dodecyl sulfate, 5 µg/ml of aprotinin, 5 µg/ml of leupeptin). Cell debris was removed by centrifugation. Protein concentration was determined with Bio-Rad protein assay. The cell lysate was boiled for 5 min in 1 x SDS sample buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1%sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol.
Our Abpromise guarantee covers the use of ab3952 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||Use at an assay dependent dilution.
HT 29 cell lysate is ready to load on SDS-PAGE for Western blotting, 10 - 20 µg per lane is recommended for mini gel.
ab3952 has not yet been referenced specifically in any publications.