Abstract
We have developed an indirect sandwich ELISA for measuring plasma apolipoprotein E (apo-E), using commercially available antibodies. A monoclonal anti-apo-E was used as the capture antibody and the captured apo-E detected with polyclonal anti-apo-E antiserum (goat). The detecting antibody was quantitated using horseradish peroxidase-conjugated rabbit immunoglobulin to goat immunoglobulins. There was no detectable cross-reactivity between the three antisera. Interference with the assay by apolipoprotein A-1, bilirubin and haemoglobin was not significant up to 1·7 g/L, 1250 μmol/L and 13·0 g/dL, respectively. The ELISA method showed high correlation with an established immunonephelometric method (ELISA apo-E=104 Immunonephelometric apo-E+16; r 2 = 0·954, P < 0·0001, n = 39). The assay has a measuring range between 5 and 560 mg/L. The coefficient of duplicates was 20%, within-run coefficients of variation (CV) ranged from 3·7 to 6·0% and between-run CV from 6·1 to 15·1%. The reference range determined for 168 normotriglyceridaemic subjects was 20 to 130 mg/L. In an analysis of the lipoprotein subfractions isolated by ultracentrifugation as the fraction of density less than 1·25 g/mL and separated by gel permeation chromatography, apo-E was found to be associated with very low-density lipoprotein and large high-density lipoprotein.