I am using TCA/acetone method to isolate proteins from plant pollen grains. When I suspend the protein pellet in 3% SDS/Tris-HCl buffer, three phases are observed after centrifugation; a pellet at the bottom, an aqueous supernatant in the middle , and a white tiny layer at the top that seems to be lipid.
I have used the aqueous supernatant in the middle to run a gel, and the quality of proteins sound good, but I suspect that some proteins might still be in the lipid phase?!
Any same experience or suggestion, I would appreciate it.
What plant species you select for isolate proteins from pollen grains ?
And if you have the control from leaves or flore éléments ?
I am trying to isolate proteins from cotton pollen. The method has been used for different plant tissues and pollen grains as well. However, the result is different plant to plant.
You can try precipitating the proteins in the upper layer using solid ammonium sulphate fractionation, centrifuge and then dialyse the solution against your buffer. Thereafter perform protein concentration assay on the dialysate.
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