I should do western blot experiment for detection of protein in liver sample .. i wash tissue with pbs then homogenize it with ripa lysis buffer and PMSF and Na3VO4 and then centrifuge it and collect supernatant and store it at -20 for about 4 months .. i wounder if my protein is degraded or not ?? Should i prepare another sample ??
Hello Duaa Adnan
If I were you, I would have stored my sample after lysis in RIPA buffer containing protease inhibitors at -80 degree C in aliquots such that each aliquot is just enough for the number of gels I would have to run at a time. This will help to avoid freeze-thaw cycles as repeated freeze-thaw would cause degradation of protein.
Storing the lysate at -20 degree C for 4 months is not recommended. I suggest you prepare another sample, but at the same time, try performing a Western on the stored sample and compare with the fresh new sample.
Best.
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