Determination of free,esterified, and total serum cholesterol using a sensitive color reaction

A method is described for the enzymatic determination of free and total cholesterol sequentially in either whole serum or the HDL fraction. Measurement of low levels of free cholesterol is facilitated by the substitution of sodium 2-hydroxy-3,5-dichlorobenzene-sulfonate for phenol in the peroxidase-catalyzed indicator reaction. Both free and total cholesterol may be measured in the same tube or separately. These alternatives may appeal to investigators who for various reasons may prefer to perform these determinations either sequentially or separately. The sequential approach is a means of conserving the relatively expensive enzymatic reagents.