A Multiplex RT-LAMP-Lateral Flow Platform For Comprehensive Dengue Serotype Detection

Abstract

Dengue virus (DENV) is a mosquito-borne RNA virus responsible for approximately 400 million infections annually and exists as four distinct serotypes (DENV-1-4). Secondary infection with a different serotype significantly increases the risk of severe dengue, making rapid serotype differentiation critical. Current gold-standard methods, such as reverse transcription polymerase chain reaction (RT-PCR), require expensive equipment and centralized laboratories, limiting accessibility in resource-limited settings. This project aims to develop a low-cost, portable diagnostic platform that combines reverse transcription loop-mediated isothermal amplification (RT-LAMP) with a multiplex lateral flow assay (LFA) for rapid detection and differentiation of all four dengue serotypes. Serotype-specific primers are being designed and optimized for RT-LAMP. A portable heating device utilizing a fatty acid phase change material (PCM) and an Mg-Fe alloy reaction was successfully constructed to provide instrument-free amplification. The LFA mechanism was also validated by successfully immobilizing the control capture probe, demonstrating effective salt-mediated immobilization, a clear colorimetric response, and successful biotin-mediated gold nanoparticle capture. Initial testing with synthetic DENV-2 DNA demonstrated successful RT-LAMP amplification. These findings establish the feasibility of an inexpensive, field-deployable RT-LAMP-LFA platform with potential to improve early dengue diagnosis and outbreak surveillance in resource-limited settings.