Isothermal Amplification

Isothermal amplification (including HAD, MCA, and LAMP) is a DNA amplification technique that has played an increasingly important role in point-of-care (POC) diagnostics. It offers several advantages over qPCR and other amplification techniques in terms of sample-to-answer time, sensitivity, specificity, cost, robustness, and accessibility, making it ideal for field-deployable diagnostics in resource-limited regions. Isothermal amplification assays have been used for numerous applications including the detection of pathogens such as salmonella and malaria, GM crop contamination, and forensics to specifically detect human DNA.

Loop mediated isothermal amplification LAMP, for example, uses four to six primers that recognize several specific regions in the target DNA. The reaction is initiated by a strand-displacement polymerase such as Bst DNA polymerase and two specially designed primers which form loop structures to facilitate subsequent rounds of amplification producing high levels of DNA. LAMP reactions are very robust and inhibitor-tolerant, producing extremely large amounts of DNA that can be visualized by the eye (e.g. as a precipitate or color change). Specifically, 1 to 10 copies of DNA can be amplified to 109 to 1010 copies within 30 minutes, producing assays with excellent sensitivity and specificity.

For even greater tolerance, Meridian has also developed sample specific inhibitor tolerant LAMP mixes, designed to be used for direct amplification from crude lysates or inhibitor-rich samples, with the ability to tolerate even higher levels of inhibitors present in samples. In addition these mixes can be used as wet mixes or dried by lyophilization or air-drying for room temperature shipping and storage, extended shelf-life, and increased flexibility in sample volume, making them ideal for POCT development where minimal sample processing and fast turn-around times are essential.


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