Are there any major protocol modifications needed? Could the high lipid content cause problems when Western blotting?
I was thinking that when you spin the tissue homogenates you can remove or avoid the fatty layer on top. Or you can perform some sort of lipid extraction (such as a Folch extraction). However I am concerned that you couild lose proteins that associate with the cellular lipids.
You didn't mention what type of protein is that. Is it extra-cellular protein or intra-cellular? There are lots of process are remaining to extract this two types of protein. If your desired protein is intra-cellular then you need to devellop a process for the cell lysis. After that you need to follow some down stream processes for the extraction of desired protein from the mixture for example Liquid chromatography or marble antibody system.
If you use a detergent such as SDS, it should get rid of the lipid problem and also allow the lipid associated proteins to be captured. However, there may be issues with solubility of the lipid associated proteins after lipid removal.
Hi Joshua, your concern is right, any manipulation of the tissue will result in losing protein. To remove lipids from protein homogenates you can use an acetone precipitation step after homogenizing the tissue.
The technique that works best for triglyceride rich lipoproteins is to remove all water with lyophilization (freeze -dry). Treat the residual with ethanol-ether (2:1) until a dry powder is left and solubilize in decyl-sulfate (not dodecyl). You may need about 0.1 M to fully solubilize but you can adjust the concentration of the decyl'sulfate by dialysis. Do-decyl and many other detergents tend to form micelles and will not dialyes easily.
Folsch extraction is too harsh for many proteins. You may need to use less ethanol and more ether depending on the solubility of the proteins in question in ethanol.
I hope that helps.
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