Phytoplankton monitoring is essential for better understanding and mitigation of phytoplankton bloom formation. We present a microfluidic cytometer with two imaging modalities for onsite detection and identification of phytoplankton: a lensless imaging mode for morphological features, and a fluorescence imaging mode for autofluorescence signal of phytoplankton. Both imaging modes are integrated in a microfluidic device with a field of view (FoV) of 3.7 mm × 2.4 mm and a depth of field (DoF) of 0.8 mm. The particles in the water flow channel can be detected and classified with automated image processing algorithms and machine learning models using their morphology and fluorescence features. The performance of the device was demonstrated by measuring Chlamydomonas, Euglena, and non-fluorescent beads in both separate and mixed flow samples. The recall rates for Chlamydomonas and Euglena ware 93.6% and 94.4%. The dual-modality imaging approach enabled observing both morphology and fluorescence features with a large DoF and FoV which contribute to high-throughput analysis. Moreover, this imaging flow cytometer platform is portable, low-cost, and shows potential in the onsite phytoplankton monitoring.

Continuous measurement of dissolved oxygen (DO) variation is important in water monitoring and biomedical applications, which require low-cost and low-maintenance sensors capable of automated operation. A frequency-domain optofluidic DO sensor with total internal reflection (TIR) design has been developed based on fluorescence quenching of Ruthenium complex (Ru(dpp) <sub xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">3</sub> Cl <sub xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">2</sub> ). To minimize artifacts causing drift in fluorescence measurements such as background autofluorescence, photobleaching, optical alignment variation, a low-cost frequency-domain approach is implemented in an optofluidic platform to measure the phase shift between the excitation and emission light. We show that the frequency domain optofluidic DO sensor provides absolute DO concentrations in repeated measurements. TIR design can enhance fluorescence signal in the integrated device and minimize background autofluorescence in the sample, which can subsequently improve overall sensitivity. Furthermore, photobleaching in the samples would be mitigated as the incident light does not enter the microfluidic channel. Our results demonstrate a measurement resolution of 0.2 ppm and response times of less than one minute. In accelerated photobleaching conditions, the long-term drift is shown to be less than ±0.4 ppm. These results suggest the potential of this optofluidic DO sensor as an in situ platform for water monitoring and biomedical applications.

The development of compact and low-cost dissolved oxygen (DO) sensors is essential for the continuous in situ monitoring of environmental water quality and wastewater treatment processes. The optical detection of dynamic and reversible quenching of fluorescent dyes by oxygen has been used for DO sensing. In this paper, we have optimized a multilayer optofluidic device based on the measurement of fluorescence quenching in a Ruthenium-based oxygen sensitive dye by employing total internal reflection (TIR) of the excitation light to achieve sensitivity enhancement for the detection of 0-20-ppm DO in water. The incident angles of light and sensitive layer thickness are optimized experimentally in order to increase the path length of light in the sensitive layer of the device through multiple reflections. A model is developed to demonstrate how light propagates through different layers of the device at varying angles of excitation and to describe the mechanism of fluorescence generation for each of the types of TIR observed. The design principles identified in this paper may be applied to the development and optimization of new multilayered optofluidic sensors by employing TIR for sensitivity enhancement.