Increasing Binding Efficiency via Reporter Shape and Flux in a Viral Nanoparticle Lateral-Flow Assay

To identify factors controlling the performance of reporter particles in a sensitive lateral-flow assay (LFA), we investigated the effect of the flux and shape of filamentous bacteriophage (phage) on the performance of phage LFAs. Phage of three different lengths and diameters were modified with biotin and AlexaFluor 555 as binding and read-out elements, respectively. The binding efficiencies of the functionalized phage were tested in a fibrous glass LFA membrane modified with avidin. The total binding rate, quantified using real-time particle counting and particle image velocimetry, decreased monotonically with the average bulk flux of phage through the membrane. At the pore scale, more phage bound in regions with faster local flow, confirming that both average and local flux increased binding. The number of bound phage increased with the aspect ratio of the phage and scaled with the phage surface area, consistent with a binding interaction controlled by the number of recognition elements on the surface. Together, these results indicate that increasing the likelihood that recognition elements on the surface of phage encounter the fibers enhances the assay binding efficiency and suggests one origin for the improved performance of nonspherical phage reporters.