Direct and Simple Fluorescence Detection Method for Oxidized Lipoproteins

Abstract

The quantification of low-density lipoprotein (LDL) and high-density lipoprotein (HDL) is currently one of the most important clinical measurements for characterizing metabolic syndrome. However, recent studies have revealed additional factors that may be more strongly associated with the coronary heart disease than simple measurement of LDL or HDL levels, such as small dense (sd) LDL particles and oxidized LDL or HDL particles. Although several methods using enzyme-antibody detection systems or fluorescent probes have been devised to characterize these factors, such methods are expensive to implement for clinical measurements. Here, we present a straightforward analytical method for direct quantitation of oxidized lipoproteins by fluorescence spectrometry, with excitation in the UV (365 +/- 10 nm) or visible (470 +/- 10 nm) range and emission detected at 450 +/- 30 nm or 535 +/- 15 nm. This method can be readily applied for clinical measurement in patients with dyslipidemia using only 1 microL of 1 mg/mL of lipoprotein and without the need for any expensive detection antibodies. Using this new technique, biological samples from patients with dyslipidemia showed higher fluorescence intensities than samples from normal subjects when detecting oxidized LDL and light HDL (d = 1.063-1.125 g/mL), whereas samples from patients with dyslipidemia showed lower fluorescence intensities than samples from normal subjects when measuring oxidized heavy HDL (d = 1.125-1.210 g/mL) levels.