Dengue is a viral infection transmitted by the bite of an infected female aedes mosquito. dengue has been recognized as one of the most important vector-borne emerging infectious diseases globally. in india, dengue has emerged as a serious public health problem. approximately 2, 93,346 people have been affected from dengue during the year of 2009-2015*, out of which 1,082 patients died. the virus of dengue is a small single stranded rna containing four distinct serotypes i.e. den1, den2, den3 and den4. these viruses belong to genus flavivirus, family flaviviridae. the dengue virus increases rapidly in human body with the increasing number of days. on first or second days of illness, the population of dengue virus is less but after 1 to 2 days ns1 starts developing and after 4 to 7 days igm and igg develop which causes severe plasma leakage leading to shock (dss) and fluid accumulation with respiratory distress. the aim of the developed paper-based device was to detect the dengue virus within one to two days from the advent of the illness so that it can be controlled from further spreading. dengue virus is diagnosed using conventional techniques like pcr (polymerase chain reaction) and elisa (enzyme-linked immunosorbent assay), but even though these methods are highly accurate they need a highly clean lab and expert people to perform these tests. they are also highly expensive and time consuming. in developing countries like india, where lab facilities are insufficient, especially in rural areas, paper-based (lateral flow assays) detection can play a complementary role. in this project, we have developed a paper-based device for diagnosis of dengue ns1 in whole blood. this device is capable of detecting very low concentrations of ns1, enabling it to detect ns1 in the first few days of the disease. it is also portable and does not require any external equipment or expertise. the device is a lateral flow test which relies on antigen–antibody interactions to detect the target of interest (ns1) in human blood. in this assay format, antibodies specific to ns1 are first bound to the surface of gold nanoparticles. then the antibody is further immobilized on the paper substrate as the capture agent. the antigen binds to the immobilized antibody, resulting in visually distinguishable lines. this change in color at the test line is due to an increase in the size of the nanoparticle when the nanoparticle-antibody conjugate binds with the ns1 antigen.
We have successfully tested Dengue NS1 antigen in solution form. We have been able to detect a concentration as low as 30 ng/ml. We are working on further reduction of the limit-of-detection. After that, we will head towards developing the paper-based device. The dimensions of the device have already been optimized using COMSOL Multiphysics simulations. The device consists of a sample pad (SP), conjugate pad (CP), test line (T) and control line (C) and an absorbent pad (A). The sample and absorbent pad will be made from cotton linter material whereas the conjugate pad will be made from glass fibre. The test and control lines will be made on a nitrocellulose membrane (NC). We have designed and developed a cassette for this device, which has been sent out for printing. We are now also working on developing an inexpensive antibody printer (similar to commercially available printers) to print the respective lines on the NC membrane.