Abstract
We present the application of wide-field time-resolved fluorescence imaging methods for the study of solvent interactions and mixing in microfluidic devices. Time-resolved imaging of fluorescence polarization anisotropy allows us to image the local viscosity of fluorescence in three dimensions in order to directly monitor solvent mixing within a microfluidic channel. This provides a viscosity image acquisition time of the order of minutes, and has been applied to a steady-state laminar flow configuration. To image dynamic fluid mixing in real-time, we demonstrate high-speed fluorescence lifetime imaging at 12.3 Hz applied to DASPI, which directly exhibits a solvent viscosity-dependant fluorescence lifetime. These two methods facilitate a high degree of quantification of microfluidic flow in 3-D and/or at high speed, providing a tool for studying fluid dynamics and for developing enhanced microfluidic assays.
Highlights
Microfluidic chip technology offers many promising applications in the fields of chemistry and biology including immunoassays, DNA amplification, DNA sequence analysis, highthroughput screening, process control, environmental monitoring and cell sorting; providing advantages such as low cost fabrication, reduced reagent consumption, high analytical throughput, improved analytical performance and enhanced portability [1,2,3]
Multidimensional fluorescence imaging (MDFI) techniques, including spectrally-resolved imaging, fluorescence lifetime imaging (FLIM), polarization-resolved imaging and fluorescence correlation spectroscopy can address these issues by providing a read-out signal that is independent of intensity, fluorophore concentration and photon path-length
We have presented two applications of time-resolved fluorescence imaging to study fluidic motion and mixing within microfluidic devices by imaging solvent viscosity
Summary
Microfluidic chip technology offers many promising applications in the fields of chemistry and biology including immunoassays, DNA amplification, DNA sequence analysis, highthroughput screening, process control, environmental monitoring and cell sorting; providing advantages such as low cost fabrication, reduced reagent consumption, high analytical throughput, improved analytical performance and enhanced portability [1,2,3]. In the work presented here we add optical sectioning to this technique, implemented in a ‘widefield’, multi-photon microscope This multiphoton microscope system multiplexes the input femtosecond laser beam to produce multiple foci, and images with a wide-field detector, such that the sample can be scanned rapidly, and produces a greater signal-to-noise whilst minimizing the non-linear photodamage associated with two-photon microscopy [11]. The calculation of the rotational mobility requires a high signal to noise ratio in the measured fluorescence decay profiles that typically results in acquisition times of up to several minutes per sectioned “image”, rendering this technique suitable for imaging steady-state fluid distributions to a high degree of quantification. We employ a fluorophore (DASPI) whose fluorescence lifetime is directly dependant on the local solvent viscosity This permits real-time imaging of dynamic fluid flow
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