I have been using a classic lysis buffer in order to extract proteins from a micro-dissected tumor tissue (300µm by 400µm). I have tried dissociating the tissue with 1) collagenase 2) lysis buffer + piston 3) lysis + sonication and haven't been able to get sufficient protein concentration for western blot analysis.
Lysis Buffer: Tris pH7.5 + 1% triton + 150mM NaCl + 5mM EDTA
If anyone has any other suggestions or comments to make, it would be greatly appreciated.
Thank you,
I'm surprised that one of those methods hasn't worked. Try resuspending in lysis buffer then doing repeated freeze thaw cycles. The ice crystal formation will pierce the cells breaking their membranes.
You can also try looking at the thermo handbook. It's pretty helpful and free.
http://find.thermofisher.com/protein-biology/protein-extraction-handbook/merch
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