13 November 2018 5 7K Report

i doubt i have a problem in my extraction procedures .. i homogenize my brain in RIPA buffer NACl 150 mM , 25mM tris , SDS 0.1%, Sdium deoxycholate 1%, Triton x 1%, edta 5mM ... i use it in 1ul /mg of tissue. 2- i homogenize for 45 stokes in glass homogenizer and shake lysate for 2 hrs at 4 degree celsius in shaker and then i centrifuge and work on supernatent...i doubt i have sth wrong with my extraction? as although i have high conc of ptn no signals of beta actin in WB ...afterthis extraction i treat my sample in ratio of 1:1 in sample buffer ? is that too much amount of sds to add ? does high number of strokes could affect my ptn .plz help if ou have experience with ripa in tissues?

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