This assay employs standard sandwich enzyme-linked immunosorbent assay (ELISA) technology to detect Dengue virus antigen NS1 in the samples. Specific antibody to dengue virus antigen NS1 was precoated in microplate wells. The test samples are incubated in the wells. The Dengue virus antigen NS1 in the sample, if presents, will bind to the immobilized antibody on the wells. Following washing, the anti-NS1 antibody-HRP conjugate is added into the tested wells. After incubation and washing procedures to remove unbound substances, this reaction is visualized by the addition of the chromogen tetramethylbenzidine (TMB). After terminating the reaction with acidic stop solution, the blue color turns yellow. What can be measured at this point is the amount of color intensity proportional to the amount of NS1 captured in the wells, and to the sample. Colorless wells appear if the dengue virus antigen is negative.
The dengue virus (DENV) is a mosquito-borne single positive-stranded RNA virus (genus Flavivirus) of the family Flaviviridae. The genome of dengue virus is about 11000 bases that codes for three structural proteins, capsid protein C, membrane protein M, envelope protein E; seven nonstructural proteins, NS1, NS2a, NS2b, NS3, NS4a, NS4b, NS5; and short non-coding regions on both the 5' and 3' ends.
Four serotypes of dengue virus have been identified and are found to be the cause of dengue fever, a severe flu-like illness. WHO estimates that 50-80 million cases of dengue fever occur worldwide each year, including a potentially deadly form of the disease called dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS). Primary dengue virus infection is characterized by elevations in specific IgM antibody levels 3 to 5 days after the onset of symptoms, which generally persists for 30 to 60 days. IgG levels also become elevated after 10 to 14 days and remain detectable for life. During secondary infection, IgM levels generally rise more slowly and reach lower levels than in primary infection, while IgG levels rise rapidly from 1 to 2 days after the onset of symptoms. Generally, primary infection patients with acute dengue infection have a higher IgM/IgG ratio. IgG levels are higher typically in the secondary infection stage. An increase in antibody titer and high IgM levels indicate acute or recent infection.