Detection Of Antihaemophilic Factor By Immunoradiometric Assays In Lymph Node And Lung Extracts

Although there have been many investigations into the site of synthesis of antihaemophilic factor, the organs and cells involved are still not clearly known and a multiorgan synthetic pathway is suspected. Extraction experiments have previously not given satisfactory results because of the presence of interfering procoagulants and the lability of factor VIII coagulant activity. Immunoradiometric assays (IRMA) for factor VIII coagulant antigen (VIII CAG) detect antigens closely related to the active coagulant site of factor VIII and are less affected by extracting media. We applied an IRMA to the detection of VIII CAG in various tissue extracts. Samples of fresh normal tissue taken at autopsy were extracted and the centrifuged supernatants tested for factor VIII CAG and factor VIII antigen by radioimmunoassays using homologous and heterologous antisera respectively. Whereas factor VIII antigen was extracted readily with isotonic saline, factor VIII CAG required 1.5M sodium chloride for optimal extraction. The possibility of VIII CAG within tissues being degraded before extraction and decay in the extracts themselves was investigated with aging and proteolytic inhibitor studies. Surprisingly, higher levels of factor VIII CAG were found in some lymph nodes and lung in comparison to liver and spleen which have been more usually considered to play some part in antihaemophilic factor biosynthesis. Factor VIII CAG in all tissue extracts was reduced by the addition of a trace of a second antibody to antihaemophilic factor different to that utilized in the IRMA.