This study was conducted to identify surface antigens of the microfilarial sheath of Litomosoides carinii which are accessible to antibodies. Rabbit antisera were raised against the soluble and insoluble fractions of purified sheaths by extracting them with a buffer containing 2-mercaptoethanol and sodium dodecylsulphate. These sera and rabbit hyperimmune sera directed against homogenates of total microfilariae, mature (i.e. microfilariae liberating) female parasites and excretory–secretory products of adult females were able to agglutinate live and formaldehyde-fixed microfilariae. When the antisera directed against sheath constituents were administered to patently infected Mastomys coucha, the microfilaraemia of these animals was rapidly reduced and remained low for a period of 2–3 weeks. Antibodies specifically binding to the microfilarial surface were immunoaffinity-purified on formaldehyde-fixed microfilariae. The antibodies react with sheath antigens of 40 and 120 kDa molecular mass which are produced by the epithelium of the distal uterus of the mature female, secreted and attached to the surface of the sheaths. A 120 kDa antigen recognized by anti-sheath surface antibodies was also detected in the excretory–secretory products of in Vitro-cultured immature female L. carinii from day 30 post-infection onwards. In the excretory–secretory products of mature adult female parasites recovered on day 130 post-infection, this 120 kDa molecule was absent. However, material reacting with the antibody was detected in the stacking gel of SDS-polyacrylamide gels. This finding may indicate that the basic units forming the 120 kDa antigen of immature adults or microfilarial sheath surface antigens occur in a highly polymerized form in the excretory–secretory products of mature female parasites.

A characteristic structural feature of Toxoplasma gondii and Neospora caninum is the presence of a triple-membrane pellicle, on the zoite stages of their complex life-cycle. Here we report the results of electron microscopic studies which show that the pellicle is made of a typical plasmalemma covered on its cytoplasmic side by a system of flattened vesicles named the inner membrane complex. Using methods described previously for the purification of pellicle and plasmalemma fractions from T. gondii, we have evaluated the same methodology for the preparation of pellicles and plasmalemma from N. caninum. The approach used involved subcellular fractionation and sucrose gradient centrifugation to prepare fractions containing pellicles. Plasmalemma was prepared by extraction of this fraction with a high salt glycerol treatment. Fractions containing membrane structures were identified by electron microscopy, and the proteins and antigens present in them were subsequently studied by SDS-PAGE and Western blotting. Electron microscopy of the pellicle fractions of N. caninum demonstrated preservation of the triple-membrane structure which is identical to that found in T. gondii. SDS-PAGE of the pellicle fractions revealed it contained several major proteins. Analyses revealed that the plasmalemma of N. caninum contained 2 abundant proteins in addition to other much lower abundance antigens detectable by monoclonal antibodies. These studies therefore report, for the first time, a detailed molecular characterization of the pellicle and plasmalemma of N. caninum.

In an attempt to identify mimotopes of the surface antigens of P. falciparum-infected erythrocytes (iRBC), antibodies were eluted from iRBC that had been treated with a pool of sera from malaria-infected individuals (IHS), and were used to screen a phage display library (PDL). After repeated panning of the PDL on immobilized antibodies, phage that selectively bound to IHS were accumulated. Of 23 randomly chosen clones that were sequenced, 13 individual sequences were detected at varying frequencies and 3 of the 13 sequences had homology with membrane proteins known to exist on iRBC. The majority of phage clones (7 out of 8 clones) selected after the 4th panning bound selectively to IgG in IHS. Specific binding of the selected phage to IgG in IHS was also confirmed using 24 IHS and 11 sera from uninfected individuals. One phage clone was the most frequently found in the sequenced clones after the 4th panning, and the binding of this clone to IgG in all IHS was greater than in any serum from uninfected individuals. A rabbit antiserum against the peptide expressed on the clone specifically recognized the surface of iRBC and resulted in iRBC haemolysis.

Previous studies using monoclonal antibody (mAb) 4C9 specific for a 32 kDa antigen (p32) of Theileria parva demonstrated expression of the antigen on the surface of the sporozoite, making it a potential antigen for sporozoite neutralization. A full-length cDNA encoding the major merozoite/piroplasm surface antigen (mMPSA) of T. parva was cloned and expressed in bacteria. The expressed product reacted strongly with mAb 4C9, demonstrating identity between the p32 and mMPSA of T. parva. Using immunoblot analysis and immunoelectron microscopy with mAb 4C9 it was shown that the mMPSA is a major antigen of the merozoite and piroplasm at the cell surface, while lower levels of antigen are expressed in the sporozoite and schizont stages. Upregulation of the mMPSA occurs at merogony and can be induced by culturing schizont-infected lymphocytes at 42 °C. Recombinant mMPSA of T. parva induced high titres of specific antibodies in cattle but failed to confer protection against a T. parva sporozoite stabilate challenge. The pre-challenge sera also failed to neutralize infectivity of sporozoites in an in vitro assay. Possible reasons for the lack of parasite neutralization in vivo and in vitro are discussed.