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A Competitive, Colorimetric ELISA for Quantification of 24(S)-Hydroxycholesterol, a Biomarker of Neurodegeneration
Dominique Wilson, Rory Olson, Mike Mullenix
Enzo Life Sciences, Farmingdale, NY 11735, USA
Sponsored by Enzo Life Sciences
Cholesterol imbalance is implicated in a variety of neurodegenerative diseases including Alzheimer’s Disease, Huntington’s Disease, and Multiple Sclerosis. Excess cholesterol is removed from the brain via the hydroxylation of C24 on cholesterol by CYP46 enzyme, a highly conserved member of the cytochrome P450 family, to the more soluble 24(S)-Hydroxycholesterol (24-OHC) which is able to cross the blood brain barrier. Levels of 24-OHC in the cerebral spinal fluid (CSF) may give insight into the homeostasis of cholesterol within the central nervous system, and may serve as a valuable biomarker for neurodegenerative disease. We have developed a competitive colorimetric ELISA for the measurement of 24-OHC that can be used in both in vivo and in vitro studies. The assay is highly specific to 24-OHC, exhibiting less than 0.02% cross-reactivity to cholesterol, 22-hydroxycholesterol, 25-hydroxycholesterol, 27-hydroxycholesterol and dehydroepiandrosterone (DHEA). The assay returns 24-OHC levels in CSF consistent with levels reported in the literature, and is also suitable for tissue homogenates and tissue culture supernatants, requiring 100 L or less of sample volume. The assay has a dynamic range from 0.78 to 100 ng/mL and a time to answer of 2 hours, making it a convenient alternative to more labor- and instrument-intensive methods such as GC-MS, LC-MS and HPLC-MS.
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