We investigated the effect of antibody and complement (C) on the fluidity of the lipid bilayer of resealed membrane ghosts by means of electron spin resonance spectroscopy. In contrast to intact red cells, resealed membrane ghosts do not rupture after treatment with C and thus C-induced changes in membrane properties can be directly ascribed to the action of C. Membranes treated with antibody alone (MA) had no modifications in the fluidity parameters. Treatment of MA with normocomplementemic rabbit serum, but not with C6-deficient rabbit serum, produced a significant increase in T‖' and a decrease in Δl and Δh, which are indicative of a more rigid structure. These modifications were absent from MA treated with the first eight C components (MAC1–8) but became apparent upon addition of C9 to the MAC1–8. Cross-linkage of the membrane proteins with glutaraldehyde increased the rigidity of the lipid bilayer. Membranes pretreated with 0.025% glutaraldehyde still responded to C with an additional increment in rigidity. However, the spin label became insensitive to C when the membranes were pretreated with 0.25% glutaraldehyde. Treatment with glutaraldehyde did not impair the development of C lesions as evaluated by the release of entrapped marker molecules. Therefore, the C-induced changes in membrane fluidity, when observed, are only the concomitant of the C lesion and not an essential component of their mechanism of production.
This work was supported by Research Grant HL 15215 from the National Institutes of Health and by the Veterans Administration.