The various complexes were prepared by mixing isolated C5b-6 in the presence of detergents or phospholipids with equimolar amounts of C7, C7 and C8, or with C7, C8 and C9 in molar ratios of 1:1:1:3. C5b-9 was also prepared by treating the SC5b-9 complex purified from inulin activated serum with 6 mM Na deoxycholate (DOC) at pH 8.1 and subjecting it to gel filtration on Sepharose 6B in the presence of DOC. This treatment resulted in selective dissociation of the S-protein, in exchange for binding of DOC by C5b-9.
Detergent-binding studies showed that C5b-6, C5b-7 and C5b-8 bind 26, 28 and 65 mole 14C-DOC per mole of complex, respectively, as measured by equilibrium dialysis. C5b-9 (prepared from SC5b-9) binds 86 mole DOC per mole of complex, measured by gel filtration in the presence of 3H-DOC. DOC binding as well as cetyl trimethyl ammonium bromide (CTAB) binding of all complexes could also be demonstrated by charge shift electrophoresis.