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Characteristics
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MCF7 cell lysate was prepared by homogenization in modified RIPA buffer (50 mM TrisHCl, pH 7.4, 1 % Triton X-100, 0.2 % sodium deoxycholate, 0.2 % sodium dodecylsulfate (SDS), 1 mM sodium ethylenediaminetetraacetate, 1 mM phenylmethyl-sulfonyl fluoride, 5 µg/mL of aprotinin, 5µg/mL of leupeptin). Cell debris was removed by centrifugation. Protein concentration was determined with Bio-Rad protein assay. The lysate was boiled for 5 min in 1 x SDS sample buffer (50 mM Tris-HCl pH 6.8, 12.5 % glycerol, 1 % SDS, 0.01 % bromophenol blue) containing 5 % Beta-mercaptoethanol.
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