Normal and homogeneous red blood cell populations over a wide range of hyper-iso-hypotonic media. III. Corrected volumes in Coulter Counter measurements.

Abstract

The volumes of human erythrocytes after rapid and gradual swelling in hypotonic NaCl media were measured using a Coulter Counter ZB at temperatures of +4 degrees C and +20 degrees C together with potassium leakage, the degree of hemolysis and the 'returning volume', i.e., the volume in an isotonic solution to which the cells will return from that in a hypotonic solution. The methodological and systematic errors in the volume measurements were corrected by taking into account the shape dependence of the Coulter Counter and the change in cell population during hemolysis, whereafter the measured cell volume values and the comparison between them become more reliable. The curves for cell volume as an inverse function of osmotic pressure appeared to be non-linear. The slopes were small at first but showed a rapid increase as the cells approached their maximal volume. The critical hemolytic volume was approx. 8% higher at +20 degrees C after both rapid and gradual swelling than at +4 degrees C and approx. 4% higher after a gradual swelling as compared with a rapid swelling both at +4 degrees C and +20 degrees C. A decrease in temperature resulted in an increase in the critical osmotic pressure both in rapid and gradual hemolysis, but did not greatly affect the amount of prelytic K+ leakage. The critical osmotic pressure was smaller in gradual hemolysis than in rapid hemolysis and the prelytic K+ leakage was doubled at both +4 degrees C and +20 degrees C. The shifts in osmotic fragility as a function of temperature may be due to differences in the visco-elastic properties of the cell membrane, but the shifts in osmotic fragility as a function of swelling rate may be connected with differences in potassium leakage and membrane stretch.