Red blood cell volume can be independently determined in vitro using sheep and human red blood cells labeled at different densities of biotin

The development of valid methods for repeatedly measuring red blood cell (RBC) volume (RCV) in the same individual would be useful in furthering understanding of the physiology and pathophysiology of the pregnant woman, fetus, and infant under a variety of conditions. Small volumes (5 to 100 mL) of either sheep or human blood were used to test the hypothesis that there is no significant difference in RCV and blood volume determined in vitro using as many as five populations of RBCs labeled at distinct biotin densities. By varying the mass of biotinylating reagent, the density of biotin on the surface of RBCs was incrementally increased to produce discrete populations as assessed by flow cytometric enumeration. Calculation of RCV for each biotin-labeled RBC population was based on the dilution principle. All biotin densities, except the most densely labeled, where variance was the greatest, accurately quantitated the in vitro blood volume to within 10 percent of the correct value. There was no bias of either overestimation or underestimation in the determination of the blood volume using either sheep or human RBCs. These in vitro results provide evidence that the multidensity biotin labeling method is sufficiently accurate to utilize in vivo for repeated determination of circulating RCV and blood volume.