MALARIA IMMUNITY AND VACCINE (UNIV HAWAII)

falciparum (PF) antigens. Large-scale production of the PF antigen will be achieved via an in vitro (IV) critical dilution cloning procedure to initiate cultures with a clone of the K+ FUP strain of PF. Isopycnic Percoll continuous gradient centrifugation will be used to enrich late schizont stages of IV cultures of PF, and variations in Percoll solution density, duration, and speed of centrifugation, and size of centrifuge tubes will be studied in order to increase yields and capacity. Three techniques to improve purification of IV-cultured PF segmenter/merozoite antigen will be studied: combining saponin lysis with a mechanical method of cell breakage; removing residual erythrocytic debris in saponin treated parasite preparation by affinity chromatography; and developing a radioactive labeling method to detect erythrocytic debris. Antigenic characteristics of merozoites obtained through purified saponin treatment will be compared with those of natural-released merozoites. This research, which will be of highest priority, will use such techniques as fractionation of soluble and solubilized merozoite preparations by 2 chromotographic and 2 electrophoretic techniques; use of hyperimmune sera from previously tested Aotus monkeys as a source of protective antibodies for the detection of PF antigens; and chemical analyses of protective antigens. Vaccination experiments will be conducted on A. trivirgatus griseimembra using long-term IV-cultured PF and particulate and immunologically-defined protective antigens. Vaccine will be studied in terms of strain specificity, and the safety for human use of synthetic adjuvants Stearoyl-MDP and CP-20,961 will be determined. Immunological research will measure humoral immune responses (via a bioassay method) as well as immunoglobulin content changes in the serum of vaccinated monkeys. Amendment of 6/81 (the basis of the above abstract) extends the project through FY83. Amendment of 6/83 accelerates research on automated IV culture systems and on the use of monoclonal antibodies for antigen isolation. (PD-AAN-057) Amendment of 3/84 extends project 3 years to permit research to characterize and purify the protective antigens from the red blood cell (RBC) stages of various species of human malaria parasites. The project will: (1) develop a large-scale (eventually fully automated) IV system for producing schizont-stage PF antigen; (2) identify, characterize, and purify potential protective antigens of blood-stage PF for testing in vaccination experiments; (3) conduct vaccination studies in A. trivirgatus using purified blood-stage PF antigens in conjunction with an appropriate adjuvant; (4) continue monoclonal antibody production and study the feasibility of using recombinant DNA technology to produce relevant antigens. (PD-AAP-823) Amendment of 8/85 accelerates RBC vaccine research, especially in the areas of genetic engineering and synthetic peptide chemistry, and initiates research into vaccine adjuvants. (PD-AAR-739)