Influence of hematocrit on the measurement of lipoproteins demonstrated by the example of lipoprotein(a)

Abstract

Background. The measurement of many parameters of human blood is usually performed in plasma or serum. Since lipoproteins or apolipoproteins, for example, are found almost exclusively in the plasma fraction after low-speed centrifugation, these parameters can be expected to be distributed in a different plasma volume depending on the hematocrit value. Therefore, the measured plasma levels might be relatively too low or too high in comparison to the whole blood concentrations in the case of abnormal hematocrit levels. The aim of our experiments was to evaluate the extent of differences between whole blood and plasma concentrations, taking as an example lipoprotein(a) [Lp(a)] in hemodialysis patients with documented decreased hematocrit values. Methods. Lp(a) was measured in plasma as well as whole blood of 15 hemodialysis patients with low hematocrit values (0.29 ± 0.02) in comparison to 11 control subjects (0.45 ± 0.04). Results. Plasma concentrations were 27% higher in patients than in controls (19.7 vs. 15.5 mg/dl). The relative difference was twice as high (59%) when measured in whole blood (13.5 vs. 8.5 mg/dl). Similar relative differences were observed when whole blood concentrations of 125 hemodialysis patients and 256 controls were calculated with the formula [Lp(a) (pasma) * (1-hematocrit)]. Conclusions. Our findings clearly demonstrate that hematocrit is a strong confounding variable of lipoprotein measurement in epidemiological studies when concentrations are measured in plasma, especially in cases of abnormal hematocrit values. Furthermore, studies investigating the longitudinal changes of lipoproteins should consider potential hematocrit changes. Lipoproteins and apolipoproteins are usually measured in plasma or serum [1]. This is convenient and can be done in frozen samples. Whole blood measurements are impossible in many cases since especially colorimetric assays are disturbed by high levels of bilirubin or hemoglobin. Handling of whole blood samples using micropipettes or automated pipetting also often disturbs the measurement due to pipette clogging. Despite the uncontested advantages of measurement in plasma or serum, the possible influence of abnormal hematocrit values in case-control studies and of fluctuating hematocrit values in longitudinal studies remains to be evaluated. During recent years several studies described an association between high lipoprotein(a) [Lp(a)] plasma concentrations and coronary heart disease [2,3]. Patients with renal disease have an increased risk for coronary heart disease [4,5] and high Lp(a) plasma concentrations [6-13; reviewed in 4]. This patient group also suffers from renal anemia with low hematocrit values. The aim of this study was, therefore, to investigate the influence of hematocrit values on the measurement of parameters distributed exclusively in the plasma fraction after low-speed centrifugation. We attempted to illustrate this question by measuring Lp(a) in hemodialysis patients. We demonstrate that Lp(a) from the whole blood of hemodialysis patients is distributed in a higher plasma volume after centrifugation due to the low hematocrit levels in these patients. Therefore, the amount and possibly also the clinical relevance of Lp(a) in these patients is underestimated in relative terms when measured as plasma instead of whole blood concentration (Fig. 1).