Preliminary observations using a multi-layer ELISA method for the detection of Entamoeba histolytica trophozoite antigens in stool samples

Abstract

A method for detecting Entamoeba histolytica trophozoite antigens in aqueous solution is described. This involves coating plastic microtitre plates with a ‘catching antibody’, specific rabbit anti- E. histolytica antibody (SRAE). After adding the test material the presence of antigen is determined using two additional heterologcus antibody layers, one of ‘developing antibody’, in this case human anti- E. histolytica immunoglobulin (HAE), which is followed by a final layer of peroxidase conjugated sheep anti-human immunoglobulin antibody (SH-HRP). The specificity and sensitivity of the assay was investigated both in the model system and using stool samples from infected patients. In the model system, the test had a sensitivity equivalent to detection of approximately one amoeba per microscope coverslip (18 mm × 18 mm). Little cross reaction was observed with other intestinal parasites common to the area of Bangladesh from which the stool samples were taken. The possible use of this method in large scale screening of stool samples and in the detection of circulating antigens is discussed.