I want to determine whether the centrifugation, separation of organic phase, & chloroform: Isopropanol: NP-40 steps are necessary in sample prep as per cell biolabs Cholesterol Assay (using the hydrogen peroxide formation, after cholesterol oxidase activity, in a colorimetric reaction).
Dear Sir. Concerning your issue about the role of chloroform, isopropanol, as well as NP-40 during the extraction of cholesterol from lysed cells. Analytical methodology has been claimed to be a source of differences found in number of studies on cholesterol content of meat and poultry products (Bragagnolo 2009). Cholesterol determination procedures in foods and other biological materials usually involve lipid extraction, separation of cholesterol from interfering components or liberation of cholesterol into the free form, and measurement of isolated cholesterol (Sweeney and Weihrauch 1976; Fenton 1992; Hwang and others 2003; Dinh and others 2008). Cholesterol can be measured by means of gravimetry, titration, colorimetry, refractometry, fluorometry, and chromatography (Sweeney and Weihrauch 1976). I think the following below links may help you in your analysis:
http://onlinelibrary.wiley.com/doi/10.1111/j.1541-4337.2011.00158.x/pdf
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2952578/
Thanks
Note, the old references to NP-40 were to a commercial product that is no longer available under that name. You should be using Igepal from Sigma. This is the new name for NP-40 as of 2 decades ago. The commerical NP-40 is now a different product.
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