Abstract:With the unique advantages of high sensitivity, equipment independence, and rapid procedure, loop-mediated isothermal amplification (LAMP) has been extensively developed to detecting various pathogens. The substances of enzyme, dNTP, and other complex organic molecules in LAMP mixture are subjected to a variety of chemical changes under room temperature, many of which are quite unacceptable in terms of storage and transportation. In order to solve this problem, we tried to develop a lyophilization technology for the protection of the LAMP mixture with the addition of trehalose as a stabilizer. The predictive effects of freeze-drying and different concentrations of trehalose (5%, 10% and 15%) in LAMP mixture during storage under temperature of 4℃, 25℃ and 37℃ on amplification activity of LAMP were analyzed systematically. The non-freeze-dried reagents without trehalose were also stored at the same temperature as controls. Our results showed that the lyophilizing LAMP reagents with 15% trehalose exhibited the greatest performance. Furthermore, at 4℃, the half-life of freeze-dried mixtures with 15% trehalose (w/v) was 112.3d, which was 6.5-fold of the control. At the temperature of 25℃, the half-life of treated reagents was 36.2d, which was 8.7-fold higher than control. Even under the temperature of 37℃, the half-life was 6.2d, which was 2-fold of the control. We report here the cost effective, simple developing procedure of a freeze-drying LAMP reagent which will promote the widespread use of LAMP in the field.
