Identification of antigens of pathogenic free-living amoebae by protein immunoblotting with rabbit immune and human sera.

Abstract

Prominent antigens of pathogenic and nonpathogenic free-living amoebae were identified by using polyclonal rabbit immune sera in immunoblot assays. The intent was to determine if prominent epitopes identified with rabbit immune sera could also be recognized by human sera. With rabbit sera, the development of immunoreactive bands was restricted to molecular masses of greater than 18.5 kDa for Naegleria, Hartmannella, and Vahlkampfia antigens. Two or more broad bands of less than 18.5 kDa were prominent features in three different Acanthamoeba species. Few cross-reactive antibodies could be detected between representative species of the three different subgroups of Acanthamoeba. Naegleria antigen was likewise serologically distinct, as were Hartmannella and Vahlkampfia antigens. The relative lack of cross-reacting antibodies between the pathogenic amoebae suggested that i would be desirable to use a panel of amoebic antigens to represent the range of serologically distinct antigens for assessing reactive antibodies in human sera. In pooled human sera (10 serum specimens per pool), the appearance of minimally reactive bands ranging from 32.5 to 106 kDa was a common feature of all six antigens. A prominent band of less than 18.5 kDa was identified in the Acanthamoeba culbertsoni antigen lane in 2 of the 10 human serum specimen pools. When sera from each of the two groups were tested individually by immunoblotting, the reaction with A. culbertsoni antigen could be associated with one individual. By using a panel of amoebic antigens, this method could prove useful in recognizing undiagnosed amoebic infections by revealing specific reactive antibodies.