As described previously [18 (link)], we utilized the Total Cholesterol and Cholesteryl Ester Colorimetric Assay Kit (Biovision; Milpitas, CA, USA) for quantification of TC (cholesterol and CE), FC and CE from plasma and peritoneal fluid. Briefly, CE, FC and TC concentrations were determined in 50 μL aliquots of sample following the kit manufacturer instructions. The absorbance was measured at 570 nm using Synergy H1MFD (Hybrid multimode) microplate reader (BioTek, VT, USA). Concentrations of TC (mg/dL) were calculated by interpolation from a standard curve.
Total cholesterol and cholesteryl ester colorimetric assay kit
Manufactured by Abcam
Sourced in United States
The Total Cholesterol and Cholesteryl Ester Colorimetric Assay Kit is a laboratory tool designed to quantitatively measure the total cholesterol and cholesteryl ester levels in samples. The kit utilizes a colorimetric method to determine the concentration of these analytes.
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4 protocols using total cholesterol and cholesteryl ester colorimetric assay kit
1
Quantitative Cholesterol Assay Protocol
2
Metabolic Profile Measurement Methods
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Glucose levels were measured using a Glucose analyzer (Glucotrend, Roche, Mannheim, Germany) according to the manufacturer’s protocol. The levels of plasma triglyceride were analyzed using a Triglyceride Quantification Colorimetric Kit (BioVision Inc., Milipitas, California, USA) according to the manufacturer’s instructions. The levels of cholesterols in the plasma were quantified by Total Cholesterol and Cholesteryl Ester Colorimetric Assay Kit (BioVision Inc., Milpitas, CA, USA) according to the manufacturer’s instructions. The levels of aspartate aminotransferase (GOT) and glutamic pyruvic transaminase (GPT) in serum were evaluated using an Alanine Aminotransferase (ALT or SGPT) Activity Colorimetric Assay Kit (BioVision Inc., Milpitas, CA, USA) and Asparate Aminotransferase (AST or SGOT) Activity Colorimetric Assay Kit (BioVision Inc., Milpitas, CA, USA) according to the manufacturer’s instructions.
3
Quantifying Plasma and Peritoneal Cholesterol
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We used the Total Cholesterol and Cholesteryl Ester Colorimetric Assay Kit from Biovision to quantify TC, FC, and CE from plasma and peritoneal fluid, as previously described (15 (link)). Following the manufacturer's instructions, concentrations were determined in 50 µl aliquots of sample. The absorbance at 570 nm was measured using a Synergy H1MFD (Hybrid multimode) microplate reader (BioTek). Interpolation from a standard curve was used to calculate TC concentrations (mg/dl).
4
Quantification of Cholesteryl Esters in Ovarian Cells
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Quantification of CE from ovarian cancer/normal cells was done using Total Cholesterol and Cholesteryl Ester Colorimetric Assay Kit (Biovision; Milpitas, CA, USA) following the manufacturer’s instructions. This kit detects total cholesterol (cholesterol and CE) in the presence of cholesterol esterase in the reaction. Briefly, 1 × 106 cells were processed for lipid extraction by adding 200 μL of chloroform: Isopropanol: NP-40 (7:11:0.1) in a micro homogenizer and centrifuged at 15,000 x g. All the liquid phase (organic phase) was transferred to a new tube and air dried at 50ºC to remove the chloroform. The samples were kept under vacuum to remove organic traces. The dried samples were then dissolved in 200 μL of cholesterol assay buffer by sonication and vortexing until homogeneous. 50 μL of the sample was used for assay of CE, free cholesterol (FC) and total cholesterol (TC) concentrations. Data was expressed as mean ± SD of against primary ovarian epithelial cells (normal controls).
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