Fluorescence spectrophotometry for COVID-19 determination in clinical swab samples

Considering the limitations of the assays currently available for the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its emerging variants, a simple and rapid method using fluorescence spectrophotometry was developed to detect coronavirus disease 2019 (COVID-19). Forty clinical swab samples were collected from the nasopharyngeal and oropharyngeal cavities of COVID-19-positive and -negative. Each sample was divided into two parts. The first part of the samples was analyzed using reverse transcription-polymerase chain reaction (RT-qPCR) as the control method to identify COVID-19-positive and -negative samples. The second part of the samples was analyzed using fluorescence spectrophotometry. Fluorescence measurements were performed at excitation and emission wavelengths ranging from 200 to 800 nm. Twenty COVID-19-positive samples and twenty COVID-19-negative samples were detected based on RT-qPCR results. The fluorescence spectrum data indicated that the COVID-19-positive and -negative samples had significantly different characteristics. All positive samples could be distinguished from negative samples by fluorescence spectrophotometry. Principal component analysis showed that COVID-19-positive samples were clustered separately from COVID-19-negative samples. The specificity and accuracy of this experiment reached 100%. Limit of detection (LOD) obtained 42.20 copies/ml (Ct value of 33.65 cycles) for E gene and 63.60 copies/ml (Ct value of 31.36 cycles) for ORF1ab gene. This identification process only required 4 min. Thus, this technique offers an efficient and accurate method to identify an individual with active SARS-CoV-2 infection and can be easily adapted for the early investigation of COVID-19, in general.