The use of multiple antigen peptides in the analysis and induction of protective immune responses against infectious diseases. four strains produced peptide-specific antibodies; however, the magnitude of the humoral response indicated strong genetic restriction between the different strains of mice. Anti-MAP antibodies acknowledged stage-specific proteins around the malaria parasites in an immunofluorescence assay. In addition, serum from hybrid BALB/cJ A/J CAF1 mice that had been immunized with the tri-epitope MAP T3-CS-T1 successfully inhibited the malaria sporozoite invasion of hepatoma cells in vitro. Spleen cells from immunized mice also showed a genetically restricted cellular immune response when stimulated with the immunogen in vitro. This study indicates that well-characterized MAPs combining solid-phase synthesis and conjugation chemistries are potent immunogens and that this approach can be utilized for the development of subunit vaccines. Malaria continues to be a major cause of mortality and morbidity in tropical areas of Africa, Asia, South America, and the South Pacific, causing an estimated 300 to 400 million new cases and more than 1.1 million deaths annually (51). The emergence of parasites that are resistant to multiple drugs and of mosquitoes that are insecticide resistant has exacerbated the problem. While a number of vaccine candidates have made it into clinical trials, few have shown great promise (20, 27, 35, 39, 42, 45, 49). These factors emphasize the need for the continued development of new malaria vaccine strategies to improve public health in areas of malaria endemicity and for visitors and short-term residents of those areas. The life cycle of the malaria parasite is usually complex; the stages in humans are morphologically and antigenically distinct and immunity tends to be stage specific (10, 22). This stage-specific gene expression actually presents an opportunity to target antigens in several stages as potential vaccine candidates. The circumsporozoite (CS) protein (5) on the surface of early sporozoites, liver-stage antigen-1 (LSA-1) (17, 23, 52), expressed when sporozoites invade liver cells, and merozoite surface protein-1 (MSP-1) (21), expressed by late liver- and blood-stage parasites, are among the handful of antigens that have been shown to have stage-specific activity 6-Bnz-cAMP sodium salt to target different developmental stages of the parasite and potentially lead to better protection. Crude antigen or attenuated malaria vaccines would be hard to produce, given the difficulty and hazards associated with mass production of parasites, the potential presence of adventitious brokers, and the possibility of side effects due to incomplete attenuation. Synthetic polypeptides as vaccine antigens provide a safer alternative to these conventional vaccine approaches. Peptide vaccines can be even more effective by focusing the host immune response on epitopes known to play a role in protective immunity and have been shown to elicit better cell-mediated immunity and to induce specific antibody responses (18, 30, 34, 38), although constructs made up of linear B-cell epitopes from malaria antigens have not always met with their expected success (2, 9, 19). Both antibody-dependent and -impartial T-cell-mediated protective immune mechanisms are operative at different stages of the parasite life cycle (4, 10), so the ideal vaccine should combine epitopes identified as strong inducers of immunity. Over the past several years, considerable progress has been made toward the development and structural design of complex polypeptides to be used as antigens. Multiple antigen peptide (MAP) conjugates provide a means to include different stage-specific peptides on one molecule, resulting in a multiepitope, multistage vaccine molecule that can potentially lead to better protection. MAPs (11, 46) offer a very attractive alternative to the conventional linear peptide approach based on a small immunologically inert core molecule of radial branching lysine residues onto which a number of 6-Bnz-cAMP sodium salt peptide antigens can be anchored. This results in a large macromolecule with a unique three-dimensional configuration which has a high molar ratio of peptide antigen to core molecule and does not require a carrier protein for elicitation of the immune response. The MAP system has already been shown to be useful in immunological studies of vaccine development in malaria and other systems (7, 28, 33, 36, 47). The construction of multiepitope malaria vaccines of defined composition has been challenging, with technical difficulties in both the synthesis and purification of product. Use of classical solid-phase synthesis methodologies in making the traditional MAP presents difficulties that often result in a highly heterogeneous product (36). In many cases it becomes impractical to obtain a reasonable amount of well-characterized product to be FASLG 6-Bnz-cAMP sodium salt evaluated as a potential vaccine candidate. We 6-Bnz-cAMP sodium salt recently overcame these difficulties and have successfully synthesized a novel multiple peptide conjugate containing immunogenic epitopes from two surface proteins (CS protein and MSP-1) 6-Bnz-cAMP sodium salt and from LSA-1 that can be used to.