sILSBio LifeSpan BioSciences, Inc. Cholesterol Assay Kit (Colorimetric) LS-K304-100 (100 Tests) • Store at -20°C Introduction CHOLESTEROL is a sterol and lipid present in the cell membranes, and is transported in the bloodstream of all animals. It is used to form cell membranes and hormones, and plays important roles in cell signaling processes. Elevated levels (hypercholesterolemia) have been associated with cardiovascular diseases such as atherosclerosis; whereas, low levels (hypocholesterolemia) may be linked to depression, cancer and cerebral hemorrhage. Simple, direct and automation-ready procedures for measuring cholesterol are very desirable. LSBio's Cholesterol Assay is based on cholesterol esterase hydrolysis of cholesterol esters to form free cholesterol and cholesterol dehydrogenase catalyzed conversion of cholesterol to cholest-4-ene-3-one, in which NAD is reduced to NADH. The optical density of the formed NADH at 340 nm is directly proportionate to the cholesterol concentration in the sample. Key Features • Sensitive and accurate. Detection limit of 5 mg/dL, linearity up to 300mg/dL cholesterol in 96-well plate assay. • Convenient. Room temperature assay. No 37°C heater is needed. • High-throughput. Can be readily automated as a high-throughput 96-well plate assay for thousands of samples per day. Applications • Direct Assays: cholesterol in serum, plasma, and other biological samples. • Pharmacology: effects of drugs on cholesterol metabolism. Materials Not Supplied Pipetting (multi-channel) devices, clear bottom 96-well plate and plate reader. Storage The kit is shipped on ice. Store all kit components at -20 °C. LifeSpan BioSciences, Inc. • 2401 Fourth Avenue, Suite 900, Seattle, WA 98121 www.LSBio.com • (206) 464-1554 • [email protected]

sILSBio LifeSpan BioSciences, Inc. Cholesterol Assay Kit (Colorimetric) LS-K304-100 (100 Tests) • Store at -20°C Assay Procedure Important: bring all reagents to room temperature prior to assay. Serum and plasma samples should be clear and free of turbidity or precipitates. If present, precipitates should be removed by filtration or centrifugation in a table centrifuge. If not assayed immediately, samples can be stored at -20 to -80°C for at least one year. 1. Standard Curve. Prepare a 10-fold diluted standard (STD) by mixing 40 pL 300 mg/dL Standard and 360 pL Assay Buffer. Further dilute standard...

sILSBio LifeSpan BioSciences, Inc. Cholesterol Assay Kit (Colorimetric) LS-K304-100 (100 Tests) • Store at -20°C Sample Data Samples were run in duplicate according to the standard procedure. The cholesterol concentrations (mg/dL) were 105 ± 3 for a human serum, 155 ± 11 for human plasma, 157 ± 2 for a bovine serum, 68 ± 2 for a rat serum, 129 ± 3 for a mouse serum, 123 ± 2 for a goat serum sample. LifeSpan BioSciences, Inc. • 2401 Fourth Avenue, Suite 900, Seattle, WA 98121 www.LSBio.com • (206) 464-1554 • [email protected]