ABSTRACT
The present paper reports research done in the following areas in the Parasitology Unit of the Department of Microbiology, National University of Singapore: determination of anti-Toxoplasma gondii antibodies in different IgG subclasses; immunoblot analysis of the antibody response to T. gondii and identification of key reactive epitopes; production and characterization of monoclonal antibodies to T. gondii; and the development of an IgM capture ELISA. IgG1 is the dominant subclass involved in the humoral response to T. gondii. IgG3 and IgG4 may be produced at low but significant levels. IgG2 production was not apparent. Immunoblot analysis of the positive sera revealed that the major antigens recognized by IgG and IgM antibodies were of MW 22, 35 and 67 kDa. Many of the bands observed in blots with positive sera were also present in blots with negative sera. The 22 kDa band was seen in majority of positive sera and was generally absent in negative sera. Anti-Toxoplasma sera also revealed IgA antibodies. Immunoblot analysis of the antigen revealed a 14 kDa band that reacted with all IgA positive sera. From a panel of monoclonals produced against T. gondii, one monoclonal, 5F3 (A) was selected for development of a monoclonal based IgM capture ELISA for detection of antibodies to 22 kDa and 41 kDa epitopes of T. gondii.
Subject(s)
Animals , Antibodies, Monoclonal/diagnosis , Antibodies, Protozoan/blood , Enzyme-Linked Immunosorbent Assay , Humans , Immunoblotting , Singapore/epidemiology , Toxoplasma/immunology , Toxoplasmosis/diagnosisABSTRACT
The HK-9 strain of Entamoeba histolytica was cultured axenically. A crude extract of the trophozoites was used as antigen in the enzyme-linked immunosorbent assay (ELISA) for the determination of antiamoebic IgG, IgM, IgA and IgG subclasses in sera of patients with E. histolytica infection. Sera from amoebiasis patients had significantly higher mean ELISA values for all classes and subclasses of immunoglobulins examined compared with healthy controls. The ELISA for IgG, IgM and IgA was positive for 76 (97.4%), 34 (43.6%) and 62 (79.5%) respectively of 78 amoebiasis sera. Analysis of IgG subclasses by ELISA showed that 76.7%, 25%, 33.3% and 66.6% of the patients were positive for antiamoebic IgG1, IgG2, IgG3 and IgG4 respectively. IgG1 and IgG4 were the dominant IgG subclasses involved in amoebiasis. The role of antiamoebic antibodies in amoebiasis is discussed.
Subject(s)
Amebiasis/immunology , Antibodies, Protozoan/analysis , Cells, Cultured , Entamoebiasis/immunology , Enzyme-Linked Immunosorbent Assay , Humans , Immunoglobulin A/analysis , Immunoglobulin G/analysis , Immunoglobulin M/analysis , Liver Abscess, Amebic/immunologyABSTRACT
The sera of 78 patients with invasive amoebiasis were tested for antiamoebic antibodies by the techniques of enzyme-linked immunosorbent assay (ELISA), indirect haemagglutination (IHA), indirect fluorescent antibody (IFA) and counterimmunoelectrophoresis (CIEP). Results showed that the ELISA compared favourably with IHA and IFA tests in terms of sensitivity and specificity. ELISA, IHA and IFA detected 97.4%, 96.2% and 98.7% of the patients respectively. CIEP was the least sensitive of the 4 serological methods with a sensitivity of 88.5%. The advantages and disadvantages of the 4 serodiagnostic procedures are discussed.