Application of Molecular Diagnostic Techniques for Viral Testing

Cobo, Fernando

Invader assays (Third Wave Technologies, Madison, WS) are based on a probe amplification method with specific recognition of particular DNA structures by cleavase, a member of the FEN-1 family of DNA polymerases. In these assays, two primers are designed which hybridize to the target sequence. Several methods can be used to detect the cleavage products, but Fluorescence Resonance Energy Transfer (FRET) probes and a second invasive cleavage reaction to detect the target-specific products is the most common technique used. FRET occurs due to the interaction between the electronic excited states of two dye molecules. FRET probes are a pair of fluorescent probes placed in close proximity. The excitation is transferred from one dye molecule to the other without emission of a photon. The acceptor fluorophore emits light which is detected in specific channels. This technology is currently available for HCV genotyping.