A red-shifted Bioluminescence Resonance Energy Transfer (BRET) biosensing system for rapid measurement of plasmin activity in human plasma.

A red-shifted Bioluminescence Resonance Energy Transfer (BRET) biosensing system for rapid measurement of plasmin activity in human plasma. Anal Chim Acta. 2020 Mar 15;1102:99-108 Authors: Weihs F, Peh A, Dacres H Abstract Proteases are key signalling molecules for many physiological processes and their dysregulation is implicated in the progression of a range of diseases. Sensitive methods to measure protease activities in complex biological samples are critical for rapid disease diagnoses. The proteolytic activity of plasmin reflects the fibrinolysis state of blood and its deregulation can indicate pathologies such as bleeding events. While Bioluminescence Resonance Energy Transfer (BRET) is a powerful and sensitive method for the detection of protease activity, the commonly applied blue-shifted BRET2 system, consisting of the Renilla luciferase Rluc2 and the large-stokes shift fluorescent protein GFP2, suffers from light absorption and light scattering in human plasma samples. To address this challenge, we developed a red-shifted BRET-based plasmin sensor by substituting BRET2 with the BRET6 system. BRET6 is composed of the red-shifted RLuc8.6 luciferase linked to the red light emitting fluorescent protein TurboFP635. The BRET6 biosensor exhibited 3-fold less light absorption in plasma samples compared to the BRET2 sensor leading to an up to a 5-fold increase in sensitivity for plasmin detection in plasma. The limits of detection ...
Source: Analytica Chimica Acta - Category: Chemistry Authors: Tags: Anal Chim Acta Source Type: research