Complementmediated-invasion-inhibition-in-the-presence-of-CPSinduced-antibodies-negatively-correlated-with-cumulative-parasitemia-during-CPS-immunizations--c

Antibodies with complement-fixing capacities (P < 0.0001). Sporozoite lysis (P = 0.017), traversal (P < 0.0001), and hepatocyte invasion inhibition (P < 0.0001) by CPS-induced antibodies were strongly enhanced in the presence of active complement.

While IgG antibodies similarly recognized homologous and heterologous sporozoites, IgM binding to heterologous sporozoites was reduced (P = 0.023). Although CPS-induced antibodies did not differ in their abilities to fix complement, lyse sporozoites, or inhibit the traversal of homologous and heterologous sporozoites, heterologous sporozoite invasion was more strongly inhibited in the presence of active complement (P = 0.008). These findings demonstrate that CPS-induced antibodies have complement-fixing activity, thereby significantly further enhancing the functional inhibition of homologous and heterologous sporozoite infectivity in vitro The combined data highlight the importance of complement as an additional immune effector mechanism in preerythrocytic immunity after whole-parasite immunization against Plasmodium hemocyanin-p-azophenylarsonate (KLH-Ar) produce anti-Ar antibodies, some of which share a cross-reactive idiotype (CRI); in general, 20 to 70% of the antihapten antibody population carries the idiotype.

Large amounts of antibody can be produced by the induction of ascitic fluids, using a 9:1 ratio of complete Freund's adjuvant (CFA) to antigen. Antibodies with the CRI can be isolated by isoelectric focusing from selected mice that have produced a high concentration of the CRI. The H chains exhibit a single homogeneous sequence through the first hypervariable region and, when isolated from a large number of individual mice, appear to be invariant in the first framework region. These findings indicate that somatic mutation is not a significant factor in the determination of framework sequences. Appearance of the CRI can be suppressed in adult A/J mice by administration of rabbit anti-idiotypic antiserum prior to immunization. Such suppressed mice produce normal concentrations of anti-Ar antibodies lacking the CRI. Anti-idiotypic antibodies produced against such antibodies failed to show cross-reactivity with anti-Ar antibodies arising in idiotypically suppressed or nonsuppressed A/J mice.

Polysucrose 400 of the assay indicates that the number of such "private" idiotypes, all present on anti-Ar antibodies of a single strain, must be extremely large; this supports a somatic mechanism for the generation of diversity. The "private" idiotypes arising in suppressed, hyperimmunized mice can be adoptively transferred into multiple, irradiated (200 R) recipients by injections of spleen cells or of cells from ascitic fluids. The use of ascitic fluids permits the rapid production of a colony of mice bearing the idiotype. This should facilitate structural studies of a variety of idiotypically different molecules sharing the same (anti-Ar) specificity, as well as studies of the mechanism of suppression.of specific monoclonal antibodies. The complex of immunoglobulin plus antigen acted as an effective immunogen in both rats and mice. Stable cell lines producing anit-human alpha 2M monoclonal antibodies to numerous antigenic sites on the alpha 2M molecule have been generated by this method.

This method has also been used to produce monoclonal antibodies to human complement components. Polysucrose 400 of immunizing mice with an immunoprecipitated product derived from conventionally produced antisera is a unique approach in that the antigen is conveniently enriched without tedious and time-consuming biochemical purification. In addition, the use of the immunoprecipitated product in conjunction with the immunoprecipitating antisera allows for rapid screening of the hybridomas in the initial stages when cell growth and maintenance is critical. This method thus further simplifies the task of obtaining a specific monoclonal antibody from a complex mixture. A final consideration is the wide availability of conventionally produced antisera to numerous proteins and other biological substances that could be used in the production of monoclonal antibodies, and consequently these antibodies could be used in more detailed studies of these same molecules.