Absence of phosphorylation-induced gelation of erythrocyte membrane skeletons: a diagnostic tool for hereditary spherocytosis.

As yet there is no single test specific for the diagnosis of hereditary spherocytosis. In the search for a specific test, a method described by Pinder et al. [14] using a cAMP-independent protein kinase extracted from normal erythrocyte membranes was used. Membrane skeletons were prepared from erythrocyte ghosts by extraction with a non-ionic detergent, i.e., Triton X-100. Upon phosphorylation with c-AMP-independent protein kinase the suspension of normal membrane skeletons set to a gelatinous mass. Membrane skeletons from patients with spherocytosis failed to show this phenomenon. In order to clarify whether this phenomenological difference can be used as a diagnostic tool for hereditary spherocytosis, a semiquantitative method of observing the gelation process was used under definite shear stress conditions. We investigated 33 patients with different hemolytic anemias (spherocytosis, hereditary elliptocytosis, hereditary stomatocytosis, homozygous beta-thalassemia and enzymopenic hemolytic anemias). With the exception of spherocytosis, all preparations of membrane skeletons showed gelation after 30-50 min. Spherocytosis membrane skeletons did not show a significant gelation even after 12 h of incubation. Thus, the failing gelation is specific for the diagnosis of hereditary spherocytosis. The "gelation assay" might be a valuable method for defining patients with hemolytic anemias due to erythrocyte membrane defects. Its molecular basis and the possible importance for the pathogenesis of spherocytosis require further investigations.