A Molecular-Based Assay for Quantifying Influenza A and B Viruses

Take a molecular detection system containing a pre-filled sample receiver, a transfer cartridge, and a test base. 

The test base has reaction tubes containing T1 and T2 primers with nick sites and influenza  A and B recognition regions, an enzyme mix, and molecular beacons with a fluorophore and quencher.

Transfer the virus-containing nasopharyngeal samples in a lysis buffer from the sample receiver to the test base.

Viral lysis releases RNA, followed by the binding of T2 primer to the target region of RNA and extension by reverse transcriptase.

The other T2 primer binds to the same RNA — displacing the initially extended strand.

The T1 primer binds to the displaced strand, and DNA polymerase extends it. A nicking enzyme cuts its site, with polymerization and strand displacement, forming double-stranded DNA.

The process continues, amplifying duplexes with single-stranded overhangs.

Molecular beacons interact with overhangs, separating the fluorophore from the quencher and producing fluorescence to quantify Influenza A and B.