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Updated: May 11, 2026
Cell-free Biochemical Fluorometric Enzymatic Assay for High-throughput Measurement of Lipid Peroxidation in High Density Lipoprotein
Published on: October 12, 2017
Standardization of a method to evaluate the antioxidant capacity of high-density lipoproteins
Elena de Juan-Franco1, Antonio Pérez, Vicent Ribas
1Department of Biochemistry, Institut de Recerca, Hospital de la Santa Creu i Sant Pau, Barcelona, Spain.
Background:
A method to evaluate the antioxidant capacity of high-density lipoprotein (HDL) was developed and standardized.
Methods:
This method measure conjugated diene (CD) formation and electrophoretic mobility of low-density lipoprotein (LDL) in agarose gels in the presence and absence of HDL. HDL was isolated from 1 mL of plasma within 24 hours and oxidation assays were performed within 6 hours. Oxidation was induced by adding CuSO4. The lag phase increase in CD kinetics and the inhibition of electrophoretic mobility were defined as the HDL antioxidant capacity.
Results:
The optimal conditions for the CD assay were 2.5 μM CuSO4, LDL at 0.1 g apoB/L, HDL at 0.1 g apoA-I/L, at 37°C and for 3h 50 min. Agarose electrophoresis at 100 V, at 4°C for 50 min was then performed immediately. CD formation variability was 21.1% for inter-assay CV and 12.7% for intra-assay CV. Electrophoretic mobility was 26.5% for inter-assay CV and 2.4% for intra-assay CV. Correlation analysis showed a significant association between the antioxidant capacity of HDL and its neutral/polar lipid ratio.
Conclusions:
The method herein described measures of the HDL antioxidant capacity in a reproducible and rapid manner that can be applied to a relatively high number of samples.
