QUANTITATIVE-ANALYSIS OF CANINE PLASMA-LIPOPROTEINS

A combined ultracentrifugation/precipitation method for the measurement of lipoprotein cholesterol concentrations was developed and validated for use with canine plasma. Very low density lipoproteins (VLDL) were isolated by flotation ultracentrifugation and low density lipoproteins (LDL) separated from high density lipoproteins (HDL) by precipitation with heparin-manganese chloride. Effective separation of these classes was confirmed by agarose gel electrophoresis of native lipoproteins and by sodium dodecyl sulphate polyacrylamide gel electrophoresis of their apolipoprotein distributions. There was trace contamination of the LDL precipitate with HDL, but this represented less than 4 and 9 per cent of the total plasma HDL in normo- and hypercholesterolaemic dogs, respectively. The intra-assay and interassay coefficients of variation for LDL- and HDL-cholesterol concentrations were between 3.3 and 6.9 per cent, and 7.2 and 9.0 per cent, respectively, for plasma cholesterol concentrations between 2.67 and 8.14 mmol/litre. The intra-assay coefficient of variation for VLDL-cholesterol was 53.8 and 18.4 per cent at plasma cholesterol concentrations of 2.67 and 8.14 mmol/litre, respectively. The inter-assay coefficient of variation for VLDL was 22.5 per cent. Storage of plasma at -20-degrees-C for between two and eight.weeks did not affect VLDL-cholesterol concentrations, but led to an increase in LDL-cholesterol and a decrease in HDL-cholesterol concentrations of approximately 10 per cent. The method described is appropriate for the measurement of lipoprotein concentrations in plasma from normo- and hypercholesterolaemic dogs, but samples should not be subjected to prolonged storage before analysis.