Microfluidics, BioMEMS, and Medical Microsystems XXII

Droplet microfluidic systems, where water-in-oil emulsion droplets can function as pico-liter-volume bioreactor, enables single-cell-resolution studies at extremely high throughput. Almost all liquid handling steps can now be conducted in droplet microfluidics format. These features make this technology extremely popular for broad ranges of microbiology and biotechnology applications, especially when the sample diversity to be screened is extremely high, in other word when the library to be screened is large. However, major challenges that prevent the even more wide-spread use of these technologies are the relatively high error rates in many droplet manipulation steps, as well as the challenges in sequentially conducting several different droplet manipulation steps often required in many biological assays of interest. Here, I present several innovations in droplet microfluidics technologies that reduce the operational errors in droplet manipulation steps as well as enable stable operations over long periods of time, and thus reducing the error rates in high-throughput multi-step screening applications.

Development of biochips enabling distortion-free imaging in the microchannel filled with a culture medium (water) is required to investigate behavior of living cells in micro and nano environments. Fluoropolymer CYTOP is a promising material as a platform of biochips for the distortion-free imaging, because the refractive index of CYTOP (1.34) is almost same as 1.33 of water. In this study, we have developed a new 3D fabrication method for CYTOP by two-photon polymerized structures using a femtosecond laser as molds, which will be used for fabrication of micro and nano environment platforms for super-resolution bioimaging.

On-chip detection of flow rates in microfluidics is critical to lab-on-chip technology. Coulometry measures the charge consumed in a redox reaction at electrode surface. In this work, constant potential coulometry at 2V was performed using a CH Instruments electrochemical workstation (Model 660E) on Titanium-Platinum microelectrodes submerged in a moving electrolyte. For a constant applied voltage, the rate of decay of current with time is observed to be a function of the flow rate (flux of ions per unit time). As the flow rate increased, the ions available at the electrode surface increased per unit time leading to slower decay rates. The model equation for current-time curve was obtained as I=αe^(-(t/τ) ); where α=0.0002 ± (1% of 0.0002) and τ=0.312 to 0.797 for electrolyte (0.1 M NaCl) flow rates ranging from 0 to 200 µL/min. The sensitivity and 3σ resolution of the flow sensor are 0.10 sec/(µL/min)/mm2 and 3.64 µL/min respectively. The sensing platform consists of micropatterned electrodes on a Si wafer plasma bonded to a PDMS microchannel on top of the electrodes. This work models the coulometric current-time curve as a first order decay problem.

Microneedles are medical microdevices that provide a pathway across the skin barrier to exchange fluids or compounds with the body for drug delivery or biosensing. This talk will highlight some of the research results on hollow microneedles and associated microfluidics phenomena including manufacturing methods for hollow microneedles. It further discusses the use of these needles to characterize the diffusion of compounds in biological tissues and the mechanics of fluid absorption biological tissue upon injection. Sensing applications of hollow microneedles include continuous glucose monitoring and a point-of-care sensing platform technology for therapeutic drug monitoring in tiny fluid volumes.

This study aimed to fabricate a droplet-based microfluidic glucose sensor using gold nanoparticles at Linoleic acid (GNPs@LA). A rapid and biocompatible method was employed to synthesize GNPs@LA. These GNPs@LA exhibit fluorescence at an excitation wavelength of 405 nm. To address the challenge of working with two immiscible solutions, namely the glucose solution and GNPs@LA, we have fabricated a droplet-based microfluidics sensor. Subsequently, the fluorescence spectra of the GNPs@LA droplets were measured. The proposed sensor utilizes the interaction between glucose and GNPs@LA, in which the fluorescence spectra's peak intensity reduces with an increase in the glucose concentration.

Fear of needles is prevalent, estimated at 63.2% in worldwide adults. We report new solid microneedle arrays made of gold in combination with micro holes to replace traditional needles for drug delivery. This work provides a breakthrough for painless drug delivery with precise control over the microneedle width, height, array density and position. Results show successful delivery of liquid drugs into the epidermal layer of porcine tissue, mimicking human skin. This scalable, fast, and automated fabrication process will enhance patient experience, improve treatment compliance, and transform drug administration into an economical and patient-friendly process at a large scale.

Flexible hearing aids can benefit from piezoelectric actuators on flexible substrates to generate vibrations on epidermis layer and bypass conductive hearing loss in a noninvasive manner. However, the major challenge is to generate a strong level of vibrations on the surface of skin that can reach cochlea with thin and low-power actuators. Lead zirconium titanate has a high piezoelectric constant and can generate vibrations with elevated levels of force and acceleration. In this paper, we assembled arrays of thin unimorph piezoelectric actuators composed of lead zirconium titanate to increase the strength of vibrations and overcome damping in flexible substrate, skin, and bone. Finite element analysis was conducted to study the displacement across substrates, skin, and bone for a single actuator and an array of actuators. Also, the experimental data show that an array of piezoelectric actuators increased the average acceleration by approximately 10 dB across 5 kHz bandwidth compared to a single actuator.

Thrombus formation is a physiological response to damage in a blood vessel. Monitoring thrombus formation is challenging, due to the limited imaging options available to analyze flowing blood. In this work, we use a visible-light optical coherence tomography (vis-OCT) system to monitor the dynamic process of the formation of thrombi in a microfluidic blood vessel-on-chip (VoC) device. Inside the VoC, thrombi form in a monolayered channel of endothelial cells perfused by human whole blood. We show that the correlation of the vis-OCT signal can be utilized as a marker for thrombus formation and can track and quantify its growth over time. We validate our results with fluorescence microscopic imaging of fibrin and platelets.

We present a microfluidic method for sorting micro/nano particles by optical radiation pressure. First, we create a narrow particle train in the middle of a microfluidic channel by laminar flow of micro/nano particles aqueous suspension. Then, we launch laser light from the side of the channel orthogonal to the flow. The particles experience optical scattering force and deviate the flowing position from the original train. The end of the channel splits into two branches. Depending on the shape, size, or color, only the particles experiencing strong optical force travel into the one of the exit branches, and the rest flows into another. To enhance the accuracy and resolution of particle sorting, we use parabolic fluid flow speed gradient in the channel.

In this study, we propose the DEP-SERS integrated system using a microfluidic device that combines metal nanostructures and microelectrodes to characterize electrophysiological and biochemical properties of biological cells. The crossover frequency which is one of dielectrophoretic properties was used to classify changes in electrical properties of cells, and cell capture rates were compared according to the structure type of the metal electrode structure applied AC power in order to properly collect cells on the nanostructure. It is expected that Raman spectroscopy using this DEP-SERS integrated system can be performed with an improved signal-to-noise ratio by collecting them right above the nanostructure.

In this study, we propose a separation method by passing a solution containing magnetic particles through microfluidic channel within a strong magnetic field. Due to the different magnetic drag forces for different Fe particle sizes, micron-sized Fe particles with almost a single diameter can be separated. This method allows low-cost production of biocompatible single-sized iron particles that can be used as the inner core of core-shell magnetic particles for various applications such as: biofunctionalized colloidal coating for biosensing, biology, and medicine applications.

Femtosecond laser direct write (fs-LDW) is a promising technique for fine 3D printing of biomaterials such as protein due to nonlinear multiphoton absorption processes facilitating microfabrication along a designated laser light path. Proteinaceous microstructures fabricated by fs-LDW are reported to retain their native protein function. Combined with submicron feature sizes, they might offer diverse biomedical or biochip applications. Here, we report the ability to 3D print from pure fluorescent protein in the colors red, blue and green via fs-LDW. Our work highlights the capabilities of fs-LDW from pure protein with a biologically highly regarded protein.

Transparent magnesium aluminate (MAS) ceramics are of high interest for industrial applications as well as for academic research due to their excellent chemical and mechanical resistance in combination with high optical transparency. However, structuring of MAS ceramics, especially on the microscale remains challenging, requiring hazardous chemicals for etching. Processes based on nanocomposites enable faster manufacturing with higher freedom of design. We have developed novel MAS nanocomposites that can be structured with high resolution and subsequently transformed into transparent polycrystalline MAS components using sintering. This new material will significantly facilitate the fabrication of microstructured components, especially for optics and photonics.

Mass production of optical and microstructured components using injection molding is an indispensable part of our modern world. However, the production of the necessary tools is costly and not very flexible, as it depends on a few very complex processes. We have developed a new method, which enables the manufacturing of structures for optical light manipulation in glass and metal by a replication process. The method successfully replicated bronze, brass, and cobalt-chromium with surface roughnesses of Rq 8 nm and microstructures in the range of 5 µm. This approach offers a cost-effective and scalable solution for high-quality molding tools, surpassing the labor-intensive and expensive machining techniques currently used.

We report a new electrochemical bio-microelectromechanical system that will enable biofluids analysis without pretreatment steps and lies in the use of an array of microelectrodes modified with bioelectronic nano-films. The multi-sensor array generates a set of complex electrochemical signals that is analyzed by using intelligent machine learning algorithms (‘chemometrics’). Here, we demonstrate the in-situ detection of: (1) the neurotransmitter dopamine in undiluted urine (a biomarker for adrenal cancer); (2) the antipsychotic clozapine in blood to provide improved schizophrenia treatment outcomes; and (3) the drug hydroxyurea in blood to enable optimized treatment of children affected with sickle cell disease.

In this work, we develop a nano-sieve device for the on-chip concentration of nanoparticles to enhance the detection sensitivity of nucleic acid targets. A sensitive CRISPR-Cas13a assay, coupled with fluorescent nanoparticles is used to recognize the viral target RNA and release a strong fluorescent signal from the nanoparticles. After the separation process, the nanospheres are injected into a nano-sieve channel for the target concentration. The nano-sieve is packed with microbeads, where the voids between these nonfunctional microbeads were used for efficiently capturing and concentrating those nanoparticles. The nanoparticles are stacked on the front end of the microbeads for 50x on-chip concentration. This technique is used to boost the fluorescence signal of low concentration nucleic acid target, a key step for the detection of various diseases such as sepsis.

We present our laboratory’s ongoing work in developing techniques based on commercially available screen-printing inks and threads to realize electronic components and multi-level circuits on clothing, where the fabric serves as the substrate for textile-based printed circuit boards. We investigate various limitations of commercially available scree-printing inks, including the physical properties of traces such as their resistance, clearance limit, resolution, ink penetration depth, flexibility, and stretchability. We also investigate the solderability of the inked traces and threads, ability to make custom components such as resistors and capacitors, durability of traces under flexing, and attachment techniques to surface mount components.

This work presents a novel method for detecting exocytotic events from dopaminergic neurons using a droplet-based microfluidic biosensor that integrates the electrochemical properties of ion beam-induced graphitic electrodes on diamond and stereolithography (SLA) 3D printing. The device combines droplet microfluidics with graphite/diamond sensors for improved cell positioning and measurement from single cells. The amperometric current intensity generated upon exciting encapsulated cells is proportional to the dopamine concentration, enabling real-time single-cell analysis. The device is versatile, time-effective, and easy to fabricate, with reasonable sensitivity, making it an ideal candidate for multiple sensing applications.

Nucleic acid testing (NAT) is currently the most sensitive method available for identifying infectious pathogens. Nevertheless, NAT-based diagnoses developed to date mostly require sophisticated infrastructures, reagents, and skilled technicians. While readily available in reference laboratories, NATs such as PCR remain inaccessible in resource-limited settings. Although extensive efforts have been undertaken toward point-of-care (POC) molecular diagnosis, a fully validated ‘sample-in-answer-out’ NAT system has not developed due to significant challenges of portability, sample preparation, and throughput. In this talk, I will discuss several low-cost field-deployable NAT devices and systems developed in our lab in the past 8 years, especially for infectious diseases in resource-limiting areas. These NAT devices could be loaded with easily-obtainable raw samples such as finger-prick blood, making diagnostic testing faster and easier for identifying pathogens like Malaria, HIV and SARS-COV-2.

A 3D printed microfluidic device for particle sorting was demonstrated using syringe-based fluid flow. Flow speeds of 192 µm/s were shown. This demonstrated an inexpensive, quick and facile fabrication approach to microfluidic particle sorting.

Calorimetry is a label-free technique to study molecular interaction and measure their enthalpy changes in solution. We have developed a microfluidic calorimetry platform that uses optical means to measure these enthalpy changes in sub-nanoliter aqueous droplets. Thermochromic liquid crystals act as optical transducers to convert temperature changes caused by the reaction heat to a shift in their reflectance spectrum. We implemented large area fiber coupled LED illumination optics and multi spot detection along the microfluidic channel to measure the temperature changes in moving microfluidic droplets. To calibrate the temperature response of our system we implemented localized laser heating of individual droplets at 1464nm and monitored the heat dissipation of those droplets in the microfluidic channel. We showed that we can observe milli-Kelvin temperature changes in droplets with millisecond time resolution.

This paper introduces an optimized and affordable optofluidic add-on device for continuous and rapid light sheet imaging of C. elegans. Platform utilizes an optofluidic chip with integrated PDMS microlens to generate a light-sheet and rapidly images worms flowing in a microchannel via a regular microscope. Validation results on control and 6-OHDA-exposed worms demonstrates the ability of platform in qualitative and quantitative imaging of neuronal degenerations at different ages. The developed affordable platform enables a conventional fluorescent microscope to offer high-content and high-throughput light-sheet images of C. elegans populations which is invaluable to aging, neurodegeneration, and drug discovery studies.

Progress achieved in the field of stem-cell technology allows the reprogramming of patient-derived cells, obtained from urine or skin biopsies, into induced pluripotent stem cells that can then be differentiated into any cell types. Within this framework, techniques, being able to accurately and non-invasively characterize cell structure, morphology, and dynamics, represent very promising approaches to identify disease-specific cell phenotypes. Consequently, we will present how a label-free optofluidic platform, based on quantitative-phase digital holographic microscopy along with various experimental developments in microfluidics, constitutes a very appealing cell imaging methodology to identify, through the measurement of biophysical properties, specific cell phenotypes.

Genome editing comprises the most promising work in 21st century genetics. Light-induced molecular surgery is the perfect tool for safe and efficient gene editing. In this work, we used the TriPleX® waveguide platform, comprised of silicon nitride and silicon oxide, to control high power visible light. Living cells are inserted into a 100 um wide microfluidic channel, then focused into a 25 um wide section in its center using sheath flows. The cells can be readily recovered at the microfluidic channel’s output with >90% survival rate. Chemically deactivated CRISPR/Cas9 molecules are activated by the laser light for safe molecular surgery. In parallel, it is possible to detect living cells flowing in the microfluidic channel. Measuring light absorption by the analyte makes it possible to detect each individual cell passing by the laser bundle, and to microscopically verify that >97% of the cells are correctly centered in the microfluidic channel. This device represents a first step to a fully integrated on-chip flow cytometer. Early results demonstrate its efficacy in cell detection and controllable exposure, paving the way to safe molecular surgery.