We are developing a method to get cholesterol binding our nano particle through hydrophobic interaction. The cholesterol we have is solved in chloroform.
An Elisa is extremely sensitive and should work. I would just dilute your cholesterol directly into water first, and make sure the chloroform concentration is low in the Elisa, ie minimally less than 1%. I would run the standard curve with the same amount of chloroform. I have not done it, but I do think it would work. I have used etoh with estrogen, derivatives and those worked.
I have tried with weird results, since organic solvents interefere with enzymatic determinations. A possibility is to add the solution to a well and let the solvent dry. But I currently use the method described in
Int J Obes . 2013 Apr;37(4):576-83.
Low doses of grape seed procyanidins reduce adiposity and improve the plasma lipid profile in hamsters.
Caimari A, del Bas JM, Crescenti A, Arola L.
The organic solvent is evaporated under N2 and lipids resuspended in an aquous buffer by sonication. The buffer used is suitable for enzymatic determination of cholesterol. If you think this approach would work for you I can send you the protocol.
Thanks for your answer Josep, We are using almost similar process in a way but because it is new for us and have not done it before so I would be grateful if you could send the protocol to me.
Thx Macia för your comment, I think ELISA will not work for us because the final concentration is to low but we ail try that in a few weeks.
Thanks again
I suggest you to try GC analysis on RTX-65 TG capillary column that allows to separate not only sterols but even diglycerides and triglycerides in a single analysis without derivatization. I used this method to determine cholesterol concentration in eggs.
If you resolve the samples in chloroform triton x100 2% , evaporate and resolve in pbs , now it is posible to determine the cholesterol concentration with a kit hope this will help
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