Methodological considerations for the accurate determination of lead in human plasma and serum.

Abstract

Studies which accurately measure plasma or serum lead (Pb) are needed to evaluate the 'biologically active' fraction of Pb in the circulation, and to clarify the role of plasma in the transportation of Pb between different compartments of the body. We evaluated several methodological aspects which influence the determination of Pb in plasma and serum. Generally, venous blood was obtained by different sampling methods (routine and ultraclean) from 3 subjects without history of Pb exposure. After centrifugation (800 g) for 10 min, the plasma or serum was analyzed by inductively coupled plasma-high-resolution mass spectrometry (ICP-MS). Several evaluations were conducted, including 1) comparison of an ultraclean serum collection method with a plasma collection method that used a commercial Vacutainer-type tube for trace metals (EDTA anticoagulant); 2) the effect of whole blood standing time prior to centrifugation on plasma or serum Pb concentration; and 3) comparison of a method using commercial heparinized Vacutainer tubes to an ultraclean plasma sampling method that utilized a low-Pb heparin anticoagulant. Plasma or serum iron (Fe) levels were also measured to evaluate hemolysis. The 3 subjects had whole blood Pb (blood-Pb) levels of 1.8, 2.0, and 2.7 micrograms/dl. Their corresponding ultraclean serum-Pb levels were 0.40%, 0.30%, and 0.48% of their whole blood-Pb levels, respectively. By comparison, the EDTA Vacutainer method plasma-Pb values were 1.7%, 1.5%, and 2.4% of whole blood-Pb, respectively. Whole blood standing (clotting) times of 15, 40, and 70 min before centrifugation resulted in increasing ultraclean serum-Pb levels of 0.21%, 0.81%, and 1.2% of whole blood-Pb (1.8 micrograms/dl), respectively. Whole blood standing time had no effect on plasma-Pb levels when heparin Vacutainers were used, or when a low-Pb heparin was used to obtain ultraclean plasma. However, plasma collected using the commercial heparin Vacutainer method contained consistently higher and more variable Pb levels than samples collected using the ultraclean plasma-Pb method. Hemolysis, when present, contributed significantly to both plasma-Pb and serum-Pb levels. In conclusion, plasma-Pb and serum-Pb levels are dependent upon methodologic processing techniques, including Pb contamination control, redistribution due to EDTA anticoagulant, hemolysis, and time dependency in sample processing. While true plasma-Pb and serum-Pb levels by any method have yet to be defined, these data provide a methodological basis from which to investigate variation in Pb partitioning between whole blood and plasma within individuals.