A nitrocellulose membrane-based ELISA for the detection of Plasmodium infections in mosquitos

A nitrocellulose (NC) membrane was evaluated as a solid-phase support for the detection of malaria-infected mosquitos using monoclonal antibodies (MAb) with a laboratory model based on Plasmodium inui and Anopheles dirus. MAbs produced against sporozoites of the N34 strain of P. inui, and selected by immunofluorescence assay and the circumsporozoite precipitin test, were used. A one-site indirect NC-ELISA that used unlabelled MAb and enzyme-labelled anti-mouse IgG was developed. Its sensitivity was about 200 sporozoites and it reliably detected one infected mosquito in a pool of 20. This indirect NC-ELISA has the advantage that it does not require direct conjugation of the MAb to an enzyme or biotin. In the direct one-site NC-ELISA, which is also reported, the relatively simple biotinylation procedure was an alternative to the enzyme- or radiolabelled MAbs. The NC-ELISAs were simple and rapid. Furthermore, the indirect NC-ELISA can be used to detect sporozoite antigen localized in various body sectors of mosquitos.