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Updated: Aug 11, 2025
Cell-free Biochemical Fluorometric Enzymatic Assay for High-throughput Measurement of Lipid Peroxidation in High Density Lipoprotein
Published on: October 12, 2017
Fully automated immunoassay for cholesterol uptake capacity to assess high-density lipoprotein function and
Katsuhiro Murakami1, Amane Harada2, Ryuji Toh3
1Central Research Laboratories, Sysmex Corporation, 4-4-4 Takatsukadai, Nishi-Ku, Kobe, 651-2271, Japan.
Insights
A new, rapid assay measures cholesterol uptake capacity (CUC), a key HDL function, offering a faster alternative to traditional methods for assessing cardiovascular health and coronary heart disease risk.
Area of Science:
- Biochemistry
- Cardiovascular Science
- Diagnostic Assays
Background:
- High-density lipoprotein (HDL) cholesterol efflux capacity (CEC) is linked to coronary heart disease (CHD) risk but requires complex, time-consuming cell-based assays.
- A cell-free assay for cholesterol uptake capacity (CUC) was previously developed as a novel, more accessible measure of HDL functionality.
- Clinical application necessitates a rapid, automated, and reproducible assay for CUC measurement.
Purpose of the Study:
- To develop and validate a rapid, automated, chemiluminescent magnetic particle immunoassay for measuring serum CUC.
- To assess the correlation of the novel CUC assay with established CEC measurements and clinical outcomes.
- To evaluate the potential of the CUC assay for cardiovascular risk stratification.
Main Methods:
- Development of a fully automated chemiluminescent magnetic particle immunoassay for serum CUC measurement.
- Assessment of CUC in response to HDL modification (myeloperoxidase oxidation, LCAT inhibition/activation).
- Correlation analysis of CUC with CEC (normalized by ApoA1) and association with coronary revascularization events.
Main Results:
- The automated assay provides high reproducibility and completes CUC measurement within 20 minutes per sample, significantly faster than CEC assays.
- CUC levels were sensitive to HDL modifications, decreasing with oxidation or LCAT inhibition and increasing with LCAT activation.
- Serum CUC correlated with CEC and was significantly associated with the need for revascularization due to recurrent coronary lesions.
Conclusions:
- A novel, rapid, and automated assay for serum CUC has been successfully developed.
- This CUC assay serves as a valuable, high-throughput tool for assessing HDL functionality and cardiovascular health.
- The assay demonstrates potential for accurate risk stratification in patients with coronary artery disease.
Abstract:
High-density lipoprotein (HDL) cholesterol efflux capacity (CEC), which is a conventional metric of HDL function, has been associated with coronary heart disease risk. However, the CEC assay requires cultured cells and takes several days to perform. We previously established a cell-free assay to evaluate cholesterol uptake capacity (CUC) as a novel measure of HDL functionality and demonstrated its utility in coronary risk stratification. To apply this concept clinically, we developed a rapid and sensitive assay system based on a chemiluminescent magnetic particle immunoassay. The system is fully automated, providing high reproducibility. Measurement of CUC in serum is completed within 20 min per sample without HDL isolation, a notably higher throughput than that of the conventional CEC assay. CUC decreased with myeloperoxidase-mediated oxidation of HDL or in the presence of N-ethylmaleimide, an inhibitor of lecithin: cholesterol acyltransferase (LCAT), whereas CUC was enhanced by the addition of recombinant LCAT. Furthermore, CUC correlated with CEC even after being normalized by ApoA1 concentration and was significantly associated with the requirement for revascularization due to the recurrence of coronary lesions. Therefore, our new assay system shows potential for the accurate measurement of CUC in serum and permits assessing cardiovascular health.
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