@article {204, title = {Molecular diagnostics in a teacup: Non-Instrumented Nucleic Acid Amplification (NINA) for rapid, low cost detection of Salmonella enterica.}, journal = {Chin Sci Bull}, volume = {58}, year = {2013}, month = {2013 Apr}, pages = {1162-1168}, abstract = {

We report on the use of a novel non-instrumented platform to enable a Loop Mediated isothermal Amplification (LAMP) based assay for Salmonella enterica. Heat energy is provided by addition of a small amount (<150 g) of boiling water, and the reaction temperature is regulated by storing latent energy at the melting temperature of a lipid-based engineered phase change material. Endpoint classification of the reaction is achieved without opening the reaction tube by observing the fluorescence of sequence-specific FRET-based assimilating probes with a simple handheld fluorometer. At or above 22{\textdegree}C ambient temperature the non-instrumented devices could maintain reactions above a threshold temperature of 61{\textdegree}C for over 90 min-significantly longer than the 60 min reaction time. Using the simple format, detection limits were less than 20 genome copies for reactions run at ambient temperatures ranging from 8 to 36{\textdegree}C. When used with a pre-enrichment step and non-instrumented DNA extraction device, trace contaminations of Salmonella in milk close to 1 CFU/mL could be reliably detected. These findings illustrate that the non- instrumented amplification approach is a simple, viable, low-cost alternative for field-based food and agricultural diagnostics or clinical applications in developing countries.

}, issn = {1001-6538}, author = {Kubota, Ryo and LaBarre, Paul and Weigl, Bernhard H and Li, Yong and Haydock, Paul and Jenkins, Daniel M} }