A Paper-Based Loop-Mediated Isothermal Amplification Assay Coupled with a Portable Heating Device for Point-of-Care Detection of Syphilis in Low-Resource Settings

Abstract

Syphilis, a sexually transmitted disease, caused by the spirochete bacterium Treponema pallidum, remains a significant global health concern. Serological methods are widely used but have limitations in early-stage infections and immunocompromised patients. This highlights the need for rapid, accurate, and cost-effective diagnostic tools to aid in syphilis management and outbreak prevention. Paper-based analytical devices have gained considerable attention in recent years for point-of-care applications due to their simplicity, low cost, and minimal equipment requirements. In this study, we have developed a paper-based loop-mediated isothermal amplification (LAMP) assay for the detection of T. pallidum DNA in clinical samples, targeting the Pol A and RNase P genes. The platform utilized a paper substrate integrated with hydroxy naphthol blue dye for visual colorimetric detection. A portable heating platform was designed to maintain the isothermal conditions necessary for amplification. The device demonstrated excellent stability when stored at 4°C and 25°C, with reproducibility significantly reduced at 37°C. Sensitivity tests revealed a detection limit of 6.4 × 10⁻⁴ ng/μL. Clinical evaluation of the paper-based LAMP assay was conducted using 52 suspected syphilis cases and 25 healthy volunteers. The assay achieved a sensitivity of 96.15% and a specificity of 100% in detecting syphilis. This portable, cost-effective device offers a promising lab-on-a-chip diagnostic solution for syphilis and can be adapted for detecting other infectious diseases. Keywords: Treponema pallidum, Syphilis, Serological identification, Loop-mediated isothermal amplification (LAMP), Paper-based assay, Portable heating platform.