Cholesterol/Cholesteryl Ester Quantitation Kit

Description

• General description: A sensitive assay for the quantitative measurement of cholesterol and/or cholesteryl ester by a colorimetric or a fluorometric method. Measurement by spectrophotometry at OD =570 nm or fluorometry at Ex/Em =535/587 nm. This assay can measure cholesterol itself, total cholesterol (cholesterol + cholesterol ester) by adding cholesterol esterase to the reaction, or only cholesteryl ester by subtracting the value of cholesterol from the total value of cholesterol plus cholesteryl esters. The fluorometric assay can detect 0.02-1 µg/well and is 4-10 fold more sensitive than colorimetric assay. Kit contains sufficient reagents for 100 tests.
• Components: Cholesterol Reaction Buffer, Cholesterol Probe, Dimethylsulfoxide, Enzyme Mix, Cholesterol Esterase, Cholesterol Standard, and a user protocol.
• Warning: Toxicity: Multiple Toxicity Values, refer to MSDS (O)
• Preparation Note: 1. Serum: The recommended volume of serum is 0.5-5 µl. Serum samples should be diluted 10-fold in Cholesterol Assay Buffer.2. Cell Lysates and Tissue Extracts: Homogenize 1 x 106 cells or 10 mg tissue in 200 µl Chloroform:Isopropanol:NP-40 (7:11:0.1) in a microhomogenizer. Centrifuge for 5-10 min at 14,000 rpm in a microcentrifuge. Avoiding the whitish solid protein layer, collect the organic phase (lower phase) and transfer it to a clean tube; air-dry at 50°C, and vacuum-dry for 30 min to remove residual chloroform. Dissolve the dried lipids in 200 µl Cholesterol Reaction Buffer by vortexing extensively for 5 min. Use 1-50 µl extracted sample per assay. The extraction procedure can be scaled up if larger amounts of sample are desired. Use 1-50 µl of extracted sample per assay. Then adjust the volume to 50 µl/well with Cholesterol Assay Buffer. For unkown samples, we suggest testing differant amounts of sample to ensure that the readings are within the linear portion of the standard curve.
• Legal Information: CALBIOCHEM is a registered trademark of Merck KGaA, Darmstadt, Germany