Osmotic fragility curves of K⁺-equilibrated RBCs. (A) The numbers next to the curves indicate the initial [K⁺], in mM, in the extracellular media used for K⁺ equilibration, as described in “Materials and methods” for study I. Open and filled paired symbols report duplicate measurements for each condition. Left shifts relative to the untreated controls indicate RBC dehydration; right shifts indicate RBC swelling. The curves represent integrals of near Gaussian distributions.^29,40 Therefore, the approximate parallelism in the observed shifts indicates that the population variation in RBC volumes did not change much in the different hydration states relative to that in the untreated controls, despite the large changes in mean cell volume. (B). Time-dependent shifts in the osmotic fragility curves of markedly dehydrated RBCs (K⁺ equilibrated in 3 mM [K⁺]). The curve at 10 minutes approximates the initial osmotic fragility pattern of the dehydrated RBCs at the start of culture. With time (4 hours and 20 hours), there is a right shift in the osmotic fragility curves, reflecting spontaneous rehydration. The progressive decrease in slope indicates increasing population variation of RBC volumes.