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The 96-kilodalton antigen as an integral membrane protein in pathogenic Entamoeba histolytica: potential differences
B E Torian1, S L Reed, B M Flores
1Department of Microbiology, Immunology and Parasitology, Louisiana State University, New Orleans 70112.
Abstract:
A surface antigen (EH-96) of Entamoeba histolytica was demonstrated to be a plasma membrane antigen by immunoprecipitation of metabolically 35S-labeled antigen from live trophozoites, Triton X-114 detergent extracts, and plasma membrane-enriched fractions prepared by concanavalin A membrane stabilization and differential centrifugation. In addition, the antigen was localized to the plasma membrane by electron microscopy with colloidal gold. Antigen from E. histolytica strains immunoprecipitated with specific immunoglobulin M (IgM) or IgG2b monoclonal antibody was identical by one-dimensional peptide mapping with N-chlorosuccinimide. Additionally, antigen from different axenically cultivated amebae was demonstrated to be identical by N-chlorosuccinimide peptide mapping, as were peptide maps of IgG and IgM monoclonal antibody-purified antigen. The 96-kilodalton (kDa) surface antigen was identified on four axenically cultivated pathogenic isolates and on three polyxenically cultivated pathogenic isolates (zymodeme II) of E. histolytica but was absent or present in lesser quantity on six nonpathogenic polyxenically cultivated isolates. The 96-kDa antigen was detected in liver abscess fluid from four patients with amebic abscesses by enzyme-linked immunosorbent assay (ELISA) and immunoprecipitation. Two-dimensional gel electrophoresis profiles of the 96-kDa antigen purified from abscess material or from polyxenically cultivated trophozoites demonstrated that the antigens were related to the 96-kDa antigen found in axenically cultivated organisms.
Insights
The 96-kilodalton (kDa) surface antigen (EH-96) of Entamoeba histolytica is located on the plasma membrane. This antigen is present in pathogenic strains and amebic liver abscess fluid, distinguishing it from nonpathogenic isolates.
Area of Science:
- Microbiology
- Immunology
- Parasitology
Background:
- Entamoeba histolytica is an enteric protozoan parasite responsible for amebiasis.
- Amebiasis is a significant global health concern, particularly in developing countries.
- Identifying specific parasitic antigens is crucial for understanding pathogenesis and developing diagnostics.
Purpose of the Study:
- To characterize the Entamoeba histolytica surface antigen EH-96.
- To determine the cellular localization of EH-96.
- To assess the presence of EH-96 in pathogenic versus nonpathogenic isolates and clinical samples.
Main Methods:
- Immunoprecipitation using metabolically labeled trophozoites and detergent extracts.
- Plasma membrane enrichment via concanavalin A stabilization and differential centrifugation.
- Electron microscopy with colloidal gold for antigen localization.
- Peptide mapping (N-chlorosuccinimide) and two-dimensional gel electrophoresis for antigen comparison.
- Enzyme-linked immunosorbent assay (ELISA) for detection in liver abscess fluid.
Main Results:
- EH-96 was confirmed as a plasma membrane antigen in Entamoeba histolytica.
- Antigenic identity was established across different strains and purification methods via peptide mapping.
- The 96-kDa antigen was predominantly found in pathogenic E. histolytica isolates (zymodeme II) and absent or reduced in nonpathogenic ones.
- EH-96 was detected in liver abscess fluid from patients with amebic abscesses.
Conclusions:
- The 96-kDa surface antigen (EH-96) is a reliable marker for pathogenic Entamoeba histolytica.
- EH-96's localization to the plasma membrane and presence in clinical samples suggest its role in parasite-host interactions.
- Further investigation of EH-96 could lead to improved diagnostic tools for amebiasis.