Analytical validation of a direct lipoprotein(a)-cholesterol assay

Accurate measurement of lipoprotein(a)-cholesterol [Lp(a)-C] may be useful in interpreting the traditional lipid panel, particularly in patients with high Lp(a). We developed and analytically validated a direct immunocapture ELISA in a Clinical Laboratory Improvement Amendments-certified laboratory for quantifying Lp(a)-C in human plasma using an apolipoprotein(a)-specific monoclonal antibody (LPA4) coupled to magnetic beads. The linearity of the assay was found to be excellent (R² = 1.00), with % bias ranging from 0.3% (upper limit of quantification) to 16.1% (lower limit of quantification) and coefficient of variation ranging from 3.4% to 16.2%. The analytical measuring range was 0.78 mg/dl to 40.0 mg/dl, and the limit of blank and limit of detection were 0.02 mg/dl and 0.05 mg/dl, respectively. The intra- and interassay coefficients of variation, determined on four quality control samples, ranged from 4.9% to 7.6% and from 12.6% to 15.0%, respectively. No interference was observed from hemolysis (up to 0.71 g/dl), bilirubin (up to 10.1 mg/dl), or triglycerides (up to 1,082 mg/dl. Lp(a)-C was stable for at least 3 months at -70 °C and through two freeze-thaw cycles. Direct Lp(a)-C correlated strongly with Lp(a) molar concentration (r = 0.93, P < 0.001). This study reports the results of the first analytical validation of a direct method to quantify Lp(a)-C, enabling standardized quantification of Lp(a)-C suitable for research studies and clinical trials; additional clinical outcome validation will be required prior to routine clinical implementation.