Disposable microfluidic micromixers for effective capture of Cryptosporidium parvum oocysts from water samples

Abstract

BackgroundProtecting drinking water supplies from pathogens such as Cryptosporidium parvum is a major concern for water utilities worldwide. The sensitivity and specificity of current detection methods are largely determined by the effectiveness of the concentration and separation methods used. The purpose of this study is to develop micromixers able to specifically isolate and concentrate Cryptosporidium, while allowing in situ analysis.ResultsIn this study, disposable microfluidic micromixers were fabricated to effectively isolate Cryptosporidium parvum oocysts from water samples, while allowing direct observation and enabling quantification of oocysts captured in the device using high quality immunofluorescence microscopy. In parallel, quantitative analysis of the capture yield was carried out by analyzing the waste from the microfluidics outlet with an Imaging Flow Cytometer. At the optimal flow rate, capture efficiencies up to 96% were achieved in spiked samples.ConclusionsScaled microfluidic isolation and detection of Cryptosporidium parvum will provide a faster and more efficient detection method for Cryptosporidium compared to other available laboratory-scale technologies.

Critical evaluation of current isolation, detection, and genotyping methods of Cryptosporidium species and future direction

“…Huang et al 172,173 reported a label-free method with microfluidic multi-angle laser scattering, which is relatively rapid (approximately 30 minutes). Efforts have also been made to sort and pre-concentrate Cryptosporidium using electrohydrodynamic concentration, 174,175 dielectrophoresismediated contact-less separation of Cryptosporidium oocysts, [176][177][178] disposable microfluidic micromixers 179 and hydrodynamic trapping-based microscopy. 178,180 Each method has its advantages: micromixers showed capture efficiency up to 96%, exceeding that of IMS, 162 hydrodynamic trappings demonstrated promise in art-effect free imaging, 179 hydrodynamic trappings showed promise in art-effect free imaging, 180 and dielectrophoretic separation can differentiate between oocysts.…”

“…Efforts have also been made to sort and pre-concentrate Cryptosporidium using electrohydrodynamic concentration, 174,175 dielectrophoresismediated contact-less separation of Cryptosporidium oocysts, [176][177][178] disposable microfluidic micromixers 179 and hydrodynamic trapping-based microscopy. 178,180 Each method has its advantages: micromixers showed capture efficiency up to 96%, exceeding that of IMS, 162 hydrodynamic trappings demonstrated promise in art-effect free imaging, 179 hydrodynamic trappings showed promise in art-effect free imaging, 180 and dielectrophoretic separation can differentiate between oocysts. [176][177][178] However, precise fluid control is necessary for optimal functioning, suggesting that these methods are unlikely to be suitable for PON testing.…”

Toward waterborne protozoa detection using sensing technologies

“…This approach was able to detect Cryptosporidium oocysts for 60 min at a maximum flow rate of 350 µL/min (5 mL could be processed in under 15 min), with a LoD of 36 oocysts/mL (Taguchi et al, 2005). Diéguez et al (2018) purposed a disposable microfluidic micromixer, which was able to specifically capture, isolate, and concentrate Cryptosporidium from water samples. This designed device was able to analyze the quantification of captured oocysts using immunofluorescence microscopy, as well as an imaging flow cytometer.…”

Size-Based Microparticle Separation Via Inertial Lift and Dean Flow in a Spiral Microchannel Device

“…Owing to the centrifugal effect, the fluid elements along the channel centerline have greater inertia and appear to move outward around a curve, producing a pressure gradient inside the channel in the radial direction (Shields IV et al, 2015) (Amini et al, 2014) . Relatively stagnant fluid near the walls recirculates inward in a fully-bounded tube due to the centrifugal pressure gradient, eventually creating two counter-rotating vortices (Diéguez et al, 2018) (Morini, 2006). Dean number K, Reynolds number Rc and the aspect ratio of the channel AR (AR=height/width) are the parameters that affect the distribution and intensity of the secondary flow in a curved channel.…”