Isolation of decay accelerating factor (DAF) by a two-step procedure and determination of its N-terminal sequence.

Decay-accelerating factor (DAF) from human red cell membranes was purified by a two-step procedure involving anion exchange and immunoaffinity chromatography. The DAF preparations were purified to homogeneity as judged by silver staining. In several experiments, the final product yields were approximately 23% of the total DAF present in the initial membrane extracts. The purified DAF retained its ability to inhibit the classical pathway C3-convertase and to reincorporate into cell membranes. An amino-terminal sequence was obtained by gas-phase sequencing. Rabbit antibodies to a synthetic peptide representing part of this sequence reacted with purified reduced membrane DAF by Western blotting and by a solid-phase immunoradiometric assay.