An ultra-red fluorescent biosensor for highly sensitive and rapid detection of biliverdin

The important role of BV in clinical diagnostics of liver-related diseases has been established in veterinary medicine. However, the sensitivity and selectivity of the current BV assays remain relatively low compromising its wider application in clinical diagnosis. Herein, we developed a rapid and sensitive BV-detecting biosensor based on a novel far-red fluorescent protein smURFP, which produced fluorescence only through specific interaction with its cofactor BV. In our study, the binding of BV to smURFP was then systematically optimized based on the structures of the smURFP + BV complex to increase the sensitivity of our biosensor. A wide linear range from 0 μM to 25 μM was obtained in both chicken and human serum. The limit of detection (LOD) and limit of quantification (LOQ) for BV was as low as 0.4 nM and 1.5 nM in human serum, and 0.4 nM and 1.2 nM in chicken serum. To our knowledge, this is the lowest LOD that has ever been reported for a BV biosensor. Our study sheds light on the biological and clinical analysis of BV.