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Color-Coding of Microchip RT-PCR Test System for SARS-CoV-2 Detection

Journal of Biosciences and Medicines 2021年第5期9卷 94-119页

作者： Rajwant K. Gill Sikander S. Gill Irina Gelimson Maxim Slyadnev Gabriel Martinez Michelle Gaines Ryan Nunley Timea Majoros Lumex Instruments Canada Mission BC Canada 1CARE Medical Diagnostics Benicia CA USA

An RT-PCR based microchip test system for the detection of SARS-CoV-2 offers pre-loaded and lyophilized reagents in the microchip. However, the 30- and 48-microwell formats of the microchip being miniaturized and performing 1.2 μl reaction, seek visual attention during sample addition. Therefore, adding colorants as color indicator in the lyophilized matrix in the microchips or adding to sample or master mix can impart not only user-friendliness to the task of liquid handling but also precision, and color-codes for easy identification of multiple kits in the layout of the microchip without compromising PCR data quality. A panel of colorants was screened for their background intensity, spectral inertness towards detection channels of AriaDNATM analyzer, interference with the reporter dyes (FAM, Cy5 and ROX), and visibility of optimal concentration in the microwell. The concentration of the colorant displaying insignificant impact on the quality of the amplification (Ct, fluorescence, and sensitivity) in comparison to no-colorant control was chosen for inclusion in the test kit. Tartrazine, Acid Red, Brilliant Blue and FAST Green colorants lyophilized with the reagents in the SARS-CoV-2 microchips were found to be stable and suitable. Storage of microchips with Fast Green colorant was tested at 40°C, 22°C, 4°C, and -20°C for 70 days and was found to be suitable and compatible with different master mixes available as liquid or lyophilized. Additionally, the microchips pre-loaded with lyophilized reagents in the presence and absence of two colorants Tartrazine and Fast Green were validated with clinical samples of SARS-COV-2. No significant impact of these colorants both intra- and inter-microchips was observed on the Ct and intensity of amplification for the tested samples in comparison to no-colorant control. The data su

关键词： SARS-CoV-2 Colorants Microchip PCR 2-plex

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