Isolation of human antibodies to factor VIII

It has been claimed that human anti-VIII:C antibodies do not form stable complexes with factor VIII and this fact has hampered in the past the isolation of such antibodies. In this study the purification of human anti-VIII:C antibodies appearing in haemophiliac patients following replacment therapy has been achieved using two different systems. In a liquid phase system, purified human factor VIII was mixed with IgG from a haemophilic patient with a high titre antibody. Specific anti-VIII:C antibodies were recovered following filtration of the antigen-antibody complexes on Biogel A-5m, dissociation of complexes at pH 3.5 and final isolation by filtration on Sephadex G-200. In a solid phase system, the same IgG fraction was specifically bound to insolubilized human factor VIII. Purified anti-VIII:C antibodies were subsequently recovered by elution of antigen-antibody complexes with magnesium chloride. The results demonstrated that stable complexes from between anti-VIII:C antibodies and either the whole factor VIII molecule, or VIII:C dissociated by previous interaction with the antibodies. It is postulated that, in vivo, similar antigen-antibody complexes may form following replacement therapy in haemophilic patients with antibody.