Abstract
A novel method for the quantification of extracellular vesicles (EVs) in biological samples has been developed combining lateral flow immunoassay with electrochemical detection based on silver enhancement. The assay uses a robust sandwich format with different antibodies targeting tetraspanins commonly expressed in EVs (CD9 and CD63), ensuring high specificity. Here, the gold nanoparticles of the test line act as nucleation sites for silver deposition when reacted with ascorbic acid, forming an electroactive signal, proportional to the concentration of EVs. Plasma-derived EVs were isolated using both precipitation and size exclusion chromatography, enabling the evaluation of the method with samples with diffetent purities. Quatification of EVs was based on coupling screen printed electrodes to the test line of the strip and subsequent anodic stripping voltammetry. The method achieved a linear detection range up to 270 EVs/μL with a detection limit of 26 EVs/μL or 11 EVs/μL, depending on the isolation method. This method demonstrates the potential of electrochemical lateral flow immunoassays (eLFIA) as an alternative strategy for sensitive EV quantification providing a proof-of-concept towards decentralized quantification of EVs in biological samples.