Heliscreener™ RNA Helicase Unwinding Assay

The Heliscreener RNA Unwinding Assay is a biochemical assay designed to measure the RNA unwinding activity of RNA helicases in the DExH/D-box family. The assay utilizes a labeled double-stranded RNA (dsRNA) substrate that is quenched when intact and emits far-red fluorescence following unwinding. The assay can be performed in kinetic (continuous monitoring) or endpoint mode. Kinetic mode is recommended for initial velocity measurements, allowing for accurate enzyme kinetics and inhibitor IC50 determinations. Alternatively, for screening, the enzyme reaction can be stopped by adding the Stop & Detect Buffer B, and the quenched reactions can be read in endpoint mode. 

How Does Heliscreener RNA Unwinding Assay Work? Fluorescence is quenched when the Reporter RNA is intact. RNA helicases unwind the Reporter RNA, releasing the far-red fluorescence (Cy5) from the quencher (Iowa Black® Quencher). A Capture RNA (green) hybridizes to the fluorescent strand, preventing it from re-annealing with the quenching strand. The Heliscreener Assay is designed specifically for high-throughput screening (HTS), with a single-addition, mix-and-read format. It is easy to integrate into automated HTS workflows, with robust detection of RNA unwinding, and compatibility with commonly used multimode plate readers. Data quality is excellent (Z’ ≥ 0.7), and the assay uses a far-red tracer to minimize interference from fluorescent compounds and light scattering.