STUDIES ON IN VITRO ANTIBODY PRODUCTION: III. PRODUCTION OF COMPLEMENT

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Abstract
Bone marrow, spleen, liver, and thymus cells of the guinea pig were incubated in Eagle"s minimal essential medium which contained glycine-2-C14. Flasks were removed from the water bath at 0, 6, 18, 24, and 48 hours and their contents were homogenized, centrifuged, and concentrated. Experiments were carried out with the concentrated supernatant samples to determine whether or not they contained proteins with properties similar to those of complement (C′) proteins. Titrations for hemolytic activity revealed that bone marrow cells produced C′2, C′3, and C′4; spleen cells synthesized C′2 and liver slices produced C′2. Liver supernatant samples also contained C′3 activity, but proteins associated with C′3 actiyity seemed to be released rather than synthesized by liver slices. We were unable to demonstrate C activity with thymus cell supernatants. Heat inactivation (56 °C for 30 minutes), as well as EDTA inhibited the hemolytic activity of supernatant samples.Further experiments showed that these supernatant proteins were fixed to antigen–antibody complexes and to aggregated gamma-globulin. The fixation was inhibited by guinea pig serum. Immunoelectrophoresis and gel-diffusion showed that proteins synthesized in vitro migrated as beta-globulins and were antigenically similar to C′ proteins.

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