Identification of Dengue Fever Markers by Dot Immunoasay

Abstract

The aim . Development of diagnostic kit for identifying markers of dengue fever at all stages of the disease. Materials and methods . In blood serum from patients with suspected dengue fever, NS1 antigen and specific IgM and IgG were detected by immune chromatography, dot immunoassay, enzyme-linked immunosorbent assay, using commercial test systems, as well as the “Dengue Spectrum” experimental kit. Results and discussion . A diagnostic kit has been developed for the detection of dengue fever markers, based on the mechanism of simultaneous differential detection of the agent NS1 protein and IgM and IgG class antibodies with the formation of specific complexes between markers from the test sample and known capture immune reagents, in a certain order, discretely fixed on a dense substrate. It was found that the effective detection of specific IgG and IgM to dengue virus can be carried out according to the scheme in which IgG is captured on the total antigen of the virus with detection using labeled anti-human IgG antibodies, and IgM is detected by capture on anti-human IgM antibodies with detection using total viral antigen. Detection of dengue virus NS1 protein can be performed using a substrate with immobilized monoclonal antibodies to NS1 and a gold immune sol bound to antibodies against NS1. This protocol of the dot analysis provides the limit for determining the recombinant analogue of the NS1 protein equal to 100 ng/ml. Comparative testing of the kit against the panel of clinical samples showed a good agreement between the results and the data obtained using imported commercial tests. The developed kit can be used for screening clinical samples, both in laboratory and in the field.