Effect of hemoglobin concentration on nucleation and polymer formation in sickle red blood cells

We have used differential polarization imaging microscopy to measure the amount and orientation of aligned sickle hemoglobin polymer in quickly deoxygenated sickle red blood cells. Images of the angular orientation of the aligned polymer at each point in the cell allowed for determination of the inclination of individual domains, providing detailed information regarding the polymerization and elongation of sickle hemoglobin polymers ex vivo. We found that the number of aligned polymer domains increased with increasing mean cell hemoglobin concentration. Sickle and holly leaf-shaped cells contained single or few domains of aligned polymer, while more compact cells such as irreversibly sickled cells contained many domains. A new class of cells was discovered by examination of images of the angular orientation of aligned polymer, which contained a single central nucleation site, with growth of polymer occurring outward in all directions in a spherulite-like domain.