Rate of Hemolytic Antibody Production by Single Cells in Vivo in Rabbits

Abstract

Abstract Published estimates of the rate of synthesis and secretion of antibody molecules by antibody-forming tissues in vitro range from 0.5 to 5000 molecules per cell per second. In the present paper we describe an experimental approach to the estimation of the absolute rate of antibody secretion per cell by undisturbed antibody-forming cells in vivo. There is evidence that most of the serum antibody appearing during the rapid rise to peak titer after a single intravenous (i. v.) injection of sheep erythrocytes into rabbits is made in the spleen. Plots of serum antibody titers typically exhibit an initial exponential rise followed by an arithmetic rise during which most of the antibody is produced and from which a constant rate of antibody production can be calculated. Rabbits immunized with a single i. v. injection of boiled sheep erythrocyte stromata were bled frequently, then sacrificed during the period of constant arithmetic antibody production and their spleen cells assayed by the liquid plaque assay technique to detect plaque-forming cells (PFC) making IgM hemolysin. Total splenic PFC were calculated based on spleen weights. The number of extrasplenic PFC was also taken into consideration and an allowance was made for PFC in the liver and blood. The rate of antibody production per milliliter in each animal was obtained from computer analysis of data for photometric 50% hemolysin units vs time and was converted to the rate of total hemolysin production based on total serum volume for each animal. This data, together with a literature value for molecules per RBC for 50% hemolysis, led to values for molecules of hemolytic antibody per PFC per second ranging from 8000 to 20,000 with a mean of 12,000 for seven rabbits. Our estimation of the protein-synthesizing machinery required for a cell to manufacture IgM antibody at this rate leads us to conclude that an average-sized plasma cell could do this if the messenger RNA is relatively stable or if gene expansion occurs to provide rapid transcription to maintain the level of messenger required.