Patent ID: 11971419
Assignee: ENZO LIFE SCIENCES, INC.
Field: Measurement (Instruments)
Classification: CPC G  C | IPC C  G

Claim 13:
14. A method of assaying for 24(S)-hydroxycholesterol in a fluid or tissue sample from a mammal; wherein the method employs anti-24(S)-hydroxycholesterol polyclonal antibodies specific to 24(S)-hydroxycholesterol that are obtained by immunizing a rabbit with 3-O-succinoyl-24(S)-hydroxycholesterol conjugated to a carrier protein through a linkage at the 3-position; the method comprising the steps of
a. noncovalently binding second antibodies to a solid phase;
b. adding the anti-24(S)-hydroxycholesterol polyclonal antibodies to the solid phase under conditions such that the second antibodies specifically bind to the anti-24(S)-hydroxycholesterol polyclonal antibodies;
c. adding the sample and a biotinylated 24(S)-hydroxycholesterol to the solid phase under conditions such that the biotinylated 24(S)-hydroxycholesterol binds to binding sites on the anti-24(S)-hydroxycholesterol polyclonal antibodies in competition with 24(S)-hydroxycholesterol in the sample;
d. adding an enzyme conjugate that is an avidin-enzyme conjugate or a streptavidin-enzyme conjugate to the solid phase under conditions such that the enzyme conjugate specifically binds to the biotinylated 24(S)-hydroxycholesterol that bound in step c;
e. adding a substrate to the enzyme in the enzyme conjugate, wherein the substrate is converted to a colored product that absorbs light at a specified wavelength in proportion to the amount of the enzyme bound to the solid phase; and
f. measuring the absorbance of light at the specified wavelength, wherein the absorbance is inversely proportional to the amount of 24(S)-hydroxycholesterol present in the sample.