Hello, I have problem to detect a 37 kDa secreted protein in cell condition media. When I precipitate protein with TCA 10% , low level of precipitated protein I get and I can not detect it by western blot.
Multiple ways have been published on enhancing detection of secreted proteins...Please consult the following reference...
Article Toward a better analysis of secreted proteins: The example o...
Mohamed,
Have you tried to immuno precipitate your protein? This may be better than using TCA and then blotting.
Hello,
Try using acetone precipitation
Add chilled 4ml of Acetone in 1ml of your conditioned media, mix gently and keep in refrigerator overnight. Next day, spin down at max speed (13000-16000 rpm) for 25-30 mins and you will get a nice pellet. check in western blot. I am convinced with acetone then TCA. If this doesn't work then you may try ammonium precipitation method.
Check it out and let me know if you need any further help.
Best regards
Pranav
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