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The current reference method in the U.S.A. for measuring in vivo population red blood cell (RBC) kinetics utilizes 51‐chromium ((51)Cr) RBC labeling for determining red cell volume (RCV), 24-hour post-transfusion RBC recovery (PTR(24)), and long-term red cell survival (RCS). Here we provide evidence supporting adoption of a method for kinetics that uses the biotin labeled RBC (BioRBC) as a superior, versatile method for both regulatory and investigational purposes. RBC kinetic analysis using biotin-labeled RBC has important methodological, analytical, and safety advantages over (51)Cr labeled-RBC. We critically review recent advances in labeling human RBC at multiple and progressively lower biotin label densities for concurrent, accurate, and sensitive determination of both autologous and allogeneic RBC population kinetics. BioRBC methods valid for RBC kinetic studies, including successful variations used by the authors, are presented along with pharmacokinetic (PK) modeling approaches for the accurate determination of RBC PK parameters in health and disease. The advantages and limitations of the BioRBC method—including its capability of determining multiple BioRBC densities simultaneously in the same individual throughout the entire RBC life span—are presented and compared with the (51)Cr method. Finally, potential applications and limitations of kinetic BioRBC determinations are discussed.

To investigate the pathophysiology of anemia and responses to red blood cell (RBC) transfusions and erythropoietin, repeated measurement of the circulating red blood cell volume (RCV) would be useful. Ovine erythropoiesis is similar to human erythropoiesis. Accordingly, a method for measuring RCV using either human or sheep RBCs labeled at different biotin densities has been previously validated in vitro. Here preclinical studies validating this method for in vivo measurement of circulating RCV in sheep are reported.For each sheep, autologous RBCs were biotinylated were at four discrete densities (12, 24, 48, and 96µg biotinylation reagent/mL RBCs). The densities were mixed and infused intravenously. Blood was sampled five times over 1 hour beginning at 4 minutes. RCV values were determined based on dilution of each population of biotinylated RBCs and by the [(14) C]cyanate method.There was no difference among RCVs measured at all densities through 16 minutes; however, by 60 minutes, RBCs biotinylated at the highest density overestimated RCV by 7.6%. RCV values increased 41% over the hour, consistent with equilibration with a pool of RBCs sequestered in the spleen. RCV by the [(14) C]cyanate method paralleled results by the biotin method but averaged 8% greater.These studies provide evidence that all four densities of biotinylated RBCs can be used in sheep to simultaneously and independently determine RCV. We speculate that the multidensity biotinylation method will be useful both for multiple simultaneous measurements and for repeated measurement of circulating RCV and blood volume in humans.

Blood-based indicators of biotin status in humans were shown to be useful tools in several clinical situations, including pregnancy. We previously validated the activity of the biotin-dependent enzyme propionyl-coenzyme A carboxylase (PCC) in lymphocytes as a sensitive and specific blood-based indicator of marginal degrees of biotin deficiency. However, the measurement of PCC activity in population studies presents substantial analytic challenges. 3-Hydroxyisovaleryl carnitine (3HIA-carnitine) increases in response to the decreased activity of the biotin-dependent enzyme methylcrotonyl-coenzyme A carboxylase and might reflect biotin status.We sought to determine whether the plasma concentration of 3HIA-carnitine increases significantly in marginal biotin deficiency.We experimentally induced marginal, asymptomatic biotin deficiency in 10 healthy adults (8 women) by having the subjects consume undenatured egg white for 28 d; biotin status was then repleted. Plasma concentrations of 3HIA-carnitine were measured on days 0, 14, 28, 35, and 50 by liquid chromatography-mass spectroscopy.The mean plasma 3HIA-carnitine concentration increased with depletion (P0.0001) and decreased with repletion (P0.0001). Plasma 3HIA-carnitine concentrations were greater than the upper limit of normal concentrations in 7 of 10 subjects by day 14 and in 9 of 10 subjects by day 28 and decreased to within normal limits in 9 of 10 subjects by day 50.These studies provide evidence that 3HIA-carnitine is an early and sensitive indicator of marginal biotin deficiency. The ease of sample collection, small sample volume requirement, and stability of 3HIA-carnitine during storage suggest that plasma 3HIA-carnitine concentration is likely to be a useful indicator of marginal biotin deficiency for larger population studies.