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The page below is a sample from the LabCE course Molecular Methods in Clinical Microbiology. Access the complete course and earn ASCLS P.A.C.E.-approved continuing education credits by subscribing online.

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Meridian's illumigene® assay for C. difficile utilizes loop-mediated isothermal DNA amplification, or LAMP.

In standard PCR, primers anneal to single strands of DNA, with polymerase enzyme replicating both strands. In the thermocycling process, denaturing, annealing, and replication occur at different temperatures.

The LAMP process is carried out at a constant temperature of approximately 63°C and relies on the strand displacing activity of DNA polymerase. In this assay, a total of six primers are utilized, targeting a conserved region of the tcd gene (the pathogenicity locus, or PaLoc). The activity of the primers and polymerase in the reaction produce strands (copies of the target area) with stem loops at either end. Replicates of the target sequence, of varying lengths, are produced in the amplification process. A byproduct of the amplification is the formation of magnesium pyrophosphate, which forms a white precipitate leading to a turbid reaction solution. The presence of turbidity indicates a positive reaction; the absence of turbidity signifies a negative reaction.
The assay requires specific instrumentation, the illumipro-10™, but the foot print is very small. The technical procedure is not difficult, and is amenable for introduction to laboratories with minimal prior experience with molecular methodologies.