Evaluation of semi-quantitative colorimetric assays based on loop-mediated isothermal amplification indicators by using image analysis

Abstract

Colorimetric assays are some of the most convenient detection methods, creating discoloration in solutions that is visible to the naked eye. However, colorimetric reactions have some limitations regarding the variability in the color perception of individuals caused by factors such as color blindness, experience, and gender. Semi-quantitative chromatic analysis has been used as an alternative method to differentiate between two colors and accurately interpret the results from a numerical value, with high confidence. Therefore, we developed and determined the optimal model between Red-Green-Blue (RGB) and Commission Internationale de l'Eclairage (CIE) Lab color spaces to establish a semi-quantitative colorimetric assay via image analysis by the ImageJ program for loop-mediated isothermal amplification (LAMP), using the dyes malachite green and phenol red. The semi-quantitative colorimetric assays using the color distance values of the CIELab color space (ΔEab) were more suitable than those using the RGB color space (ΔERGB) for chromatic differentiation between positive and negative reactions in both indicator dyes, demonstrating the feasibility of this assay to be applied in the detection of a wide range of pathogens and infectious diseases.