Ektacytometric measurement of sickle cell deformability as a continuous function of oxygen tension

In an effort to study the rheologic effects of small amounts of hemoglobin S (HbS) polymer in sickle red cells, we have used the ektacytometer, a laser diffraction couette viscometer, to measure sickle cell deformability as a function of oxygen tension. Sickle cell populations of defined intracellular hemoglobin concentration (MCHC) were isolated using Stractan density gradients and were resuspended in buffered polyvinylpyrrolidone solutions for deformability measurements. Using a gas-porous, hollow fiber gas exchange system to establish a linear gradient in oxygen tension, deformability was measured over a pO2 range of 76 to 0 mm Hg. Parallel spectroscopic determinations of oxygen saturation permitted determination of cell deformability as a function of oxygen saturation for each discrete MCHC population. From these measurements the level of oxygen saturation at which a loss in cell deformability was first detected could be defined. Then, using the data of Noguchi and Schecter, the amount of polymerized HbS in the cells at that defined level of oxygen saturation was estimated. The results of this analysis suggested that the quantity of polymer that caused a detectable loss in cell deformability increased with increasing MCHC. In addition, for MCHC above 30 g/dL, this represented a substantial fraction of the total HbS in the cell.