End-point detection of loop-mediated isothermal amplification (LAMP) on malaria by direct observation with colorimetric dyes

In order to ascertain the results of the LAMP technique, different end-point detection methods can be employed. However, these methods require sophisticated equipment. To simplify current end-point detection methods for the diagnosis of malaria, we propose the incorporation of colorimetric dyes: malachite green (MG), phenol red (PR), and xylenol orange (XO) in the LAMP assay. To evaluate the optimum concentration of dyes, 5 different concentrations (50 mu M, 75 mu M, 100 mu M, 125 mu M, and 150 mu M) were used with buffer pH 8.5 and pH 8.8, respectively. The results showed that 125 mu M of MG at pH 8.8 produced the most obvious colour change. A total of 71 clinical blood samples of Plasmodium knowlesi, Plasmodium malariae, Plasmodium vivax, Plasmodium falciparum, and healthy donors were tested using MG -LAMP. It showed 100% sensitivity and specificity. The simplicity and affordability of this method make it ideal to be used as an end-point detection method for malaria diagnosis in resource limited settings.

Automated summary

This study proposes the use of colorimetric dyes (malachite green, phenol red, and xylenol orange) in the LAMP assay to simplify current end-point detection methods. The optimal concentration of dyes was determined to be 125μM MG at pH 8.8, which produced the most obvious color change. MG-LAMP showed 100% sensitivity and specificity when tested on 71 clinical blood samples. This method is simple, affordable, and suitable for malaria diagnosis in resource-limited settings.