In situ Tracking of Exoenzyme Activity Using Droplet Luminescence Concentrators for Ratiometric Detection of Bacteria

Abstract

We demonstrate a novel, rapid, and cost-effective biosensing paradigm that is based on an in situ visualization of bacterial exoenzyme activity using biphasic Janus emulsion droplets. Sensitization of the droplets toward dominant extracellular enzymes of bacterial pathogens is realized via selective functionalization of one hemisphere of Janus droplets with enzyme-cleavable surfactants. Surfactant cleavage results in an interfacial tension increase at the respective droplet interface, which readily transduces into a microscopically detectable change of the internal droplet morphologies. A macroscopic fluorescence read-out of such morphological transitions is obtained via ratiometrically recording the angle-dependent anisotropic emission signatures of perylene-containing droplets from two different angles. The optical read-out method facilitates detection of marginal morphological responses of polydisperse droplet samples that can be easily produced in any environment. The performance of Janus droplets as powerful optical transducers and signal amplifiers is highlighted by rapid (<4 h) and cost-effective antibody and DNA-free identification of three major foodborne pathogens, with detection thresholds of below 10 CFU mL –1 for Salmonella and <10 2 to 10 3 CFU mL –1 for Listeria and Escherichia coli .

Real-time detection and classification of PFAS using dynamic behaviors at liquid–liquid interfaces

“…We measured the real-time changes in interfacial tensions, induced by exposure to PFAS solutions, by measuring the changes in the fluorescent emission intensity from a singlelayer array of Janus droplets. This coupled relationship among the chemical properties (interfacial tensions), the physical properties (droplet morphology), and optical properties (fluorescent emission) was first reported by Swager and coworkers 56 and has been used by our group 17,18,61 and others 55,[57][58][59][60][67][68][69] as the sensing mechanism for a variety of analytes. Briefly, the direction (and measured intensity) of the emissive light of a fluorophore (perylene) is affected by the internal morphology of Janus droplets.…”

“…61,70 This sensing platform provides a rapid and continuous measurement of fluorescent emission intensity from Janus droplets, which is directly correlated to the interfacial tension and the concentration of the targeted analyte. 17,18,56,57,60,67,68 For each experiment, we collected the emission intensity as a function of time at a characteristic wavelength of perylene (λ = 475 nm) as the Janus droplets were exposed to PFAS solutions. The duration and concentration of PFAS exposure were adjusted by tuning the ratio of the flowrates of the two syringe pumps upstream from the Janus droplets, containing the control solution and PFAS solution.…”

“…Unlike conventional tensiometers that offer insights solely into equilibrium interfacial tensions at a single interface, Janus droplets are capable of monitoring dynamic interfacial tensions across multiple interfaces simultaneously and can differentiate between hydrocarbon and fluorocarbon surfactant. 18 When incorporated with responsive surfactants that are selective to targeted analytes, these droplets have been tailored to detect enzymes, 53,54 foodborne pathogens, [55][56][57] dissolved metals in water, 17,58,59 and trace chemicals. 60 We adopted an optical detection technique known as "directional emission" that correlates interfacial tensions to the fluorescent emission of Janus droplets.…”