Immunochemical characterisation of a 29-Kda surface-associated molecule of Entamoeba histolytica and its recognition by serum from patients with amoebiasis

Abstract

Summary.A 29-Kda cytotoxic molecule of axenically-grown pathogenic Entamoeba histolytica (strain HM 1) was purified from an amoebic extract by immuno-affinity chromatography with monoclonal antibodies. Immunoreactivity of the purified 29-Kda molecule altered significantly (p < 0.01) after exposure to heat or trypsin, but remained unaltered after treatment with sodium metaperiodate. The 29-Kda molecule was recognised by serum from each of 13 patients with amoebic liver abscess. In an ELISA system, the molecule produced significantly higher (p < 0.01) OD readings with these serum samples than with samples from asymptomatic cyst passers. No serum from healthy subjects or from patients with idiopathic ulcerative colitis or giardiasis had antibodies that reacted with the 29-Kda molecule. The immune response to the 29-Kda amoebic protein in man may indicate a specific role for this molecule in invasive amoebiasis.

Partial characterization of a 36-kDa antigen ofEntamoeba histolyticaand its recognition by sera from patients with amoebiasis

“…Characterization of 36‐kDa antigen was achieved by measuring its immunoreactivity to MoAb 3D 10 following exposure to (a) heat, (b) trypsin and (c) sodium metaperiodate in a micro‐ELISA system [14]. The sensitivity to heat was determined by exposing this 36‐kDa antigen to 60 or 100°C (boiling) for 10 min before coating on to the wells of the ELISA plate.…”

“…Each serum sample was tested against the 36‐kDa antigen in an ELISA system [14]. Briefly, ELISA was performed in 96‐well microtitration plates (Costar Corporation, USA) by coating wells with 100 μl of optimally diluted partially purified 36‐kDa antigen (2.0 μg ml −1 ) in 0.05 M carbonate buffer, pH 9.6.…”

Partial characterization of a 36-kDa antigen of Entamoeba histolytica and its recognition by sera from patients with amoebiasis

“…Characterization of 36-kDa antigen was achieved by measuring its immunoreactivity to MoAb 3D 10 following exposure to (a) heat, (b) trypsin and (c) sodium metaperiodate in a micro-ELISA system [14]. The sensitivity to heat was determined by exposing this 36-kDa antigen to 60 or 100³C (boiling) for 10 min before coating on to the wells of the ELISA plate.…”

“…Each serum sample was tested against the 36-kDa antigen in an ELISA system [14]. Brie£y, ELISA was performed in 96-well microtitration plates (Costar Corporation, USA) by coating wells with 100 Wl of optimally diluted partially puri¢ed 36-kDa antigen (2.0 Wg ml 31 ) in 0.05 M carbonate bu¡er, pH 9.6.…”

“…Thus, surface-associated molecules and various cytotoxic molecules are important in mediating the host parasite interactions. However, studies of the individual antigens and their biochemical and immunological properties are lack-ing, with the exception of a few, like the 220-kDa protein that is inhibited by N-acetyl-D-glucosamine [10], N-acetyl galactosamine inhibitable adherence lectin [11], a 125-kDa surface antigen [12], 96-kDa protein [13] and a 29-kDa surface-associated protein [14].…”