enhancing rapid detection of covid-19: optimization of rt-lamp assays for improved sensitivity and specificity

Background: the covid-19 pandemic has underscored the critical need for rapid and reliable diagnostic methods to detect the sars-cov-2 virus and effectively curb its transmission. traditional diagnostic methods, such as rt-pcr, although accurate, often require extensive laboratory infrastructure and time, which can hinder timely public health interventions. objectives: this study aimed to optimize a reverse transcription loop-mediated isothermal amplification (rt-lamp) assay for the direct detection of viral rna from nasopharyngeal swabs, eliminating the need for nucleic acid extraction and reducing reliance on sophisticated laboratory equipment. methods: the optimization process involved several key steps, including adjustments in sample preparation techniques, fine-tuning the concentration of proteinase k, and incorporating guanidine hydrochloride and betaine to enhance the assay's sensitivity. results: the optimized rt-lamp assay achieved a sensitivity of approximately 85.25% and a specificity of 100% when utilizing extracted rna. in comparison, the direct use of pharyngeal swab samples yielded a sensitivity of 83.61% and a specificity of 86.67%. the assay successfully identified viral rna with cycle threshold (ct) values up to 35, indicating its capability to detect low viral loads comparable to those identified by standard rt-pcr methods. test positivity cut-offs were based on a colorimetric shift and confirmed by gel electrophoresis. conclusions: this rt-lamp assay presents a promising alternative to rt-pcr for rapid covid-19 diagnostics, particularly in resource-limited settings. by providing a balance of efficiency, accuracy, and accessibility, it can facilitate timely public health responses and enhance the capacity for widespread testing during pandemics.