Advanced Lyophilised Loop Mediated Isothermal Amplification (L-LAMP) based point of care technique for the detection of dengue virus

Abstract

Dengue virus infects millions of the people globally each year and its diagnosis remains a challenge. Conventionally used diagnostic methods are complex and time consuming. LAMP technique is a potential alternative for diagnosis of dengue virus. The benefits of LAMP are its ease and ability, as it does not require an expensive equipment and results are effortlessly visualized by the naked eye. However, it does not aid as point of care technique owing to need of contamination free area, deep freezer for chemical storage and primer self amplification. Each small modification in LAMP method bring it towards an ideal point of care technique. An advanced lyophilized loop mediated isothermal amplification (L-LAMP) was developed in which the dye was dried on the cap and reaction reagents was lyophilized at the bottom of the tube to overcome the common hurdles of LAMP technique. The technique was able to diagnose disease within 35 min with 4U of Bst polymerase. The least concentration of dye required was 1000×.Result given by the seminested reverse transcriptase polymerase chain reaction (RT-PCR) and L-LAMP with enzyme linked immuno sorbent assay (ELISA) were compared using Chi square test. The L-LAMP showed 100 % specificity and 92 % sensitivity with respect ELISA and was found better than RT-PCR which showed 100 % specificity and 88 % sensitivity. There was no cross reactivity of primers with other disease like malaria caused by Plasmodium falciparum and P. vivax and with viral disease chikungunya. L-LAMP has dynamic potential as point of care technique.