Microfluidic Fabrication Research Articles

Using 3D printing to fabricate microfluidic chips for biosensing applications

This paper gives details on the use of 3D smart printing technology to fabricate microfluidic chips for integration into biosensors for the detection and diagnosis of diseases. Additive manufacturing, also known as 3D printing, is a process used to create complex, three-dimensional objects by adding layer upon layer of material until the desired shape is formed. Microfluidic chips are used to manipulate fluids through separation and mixing. Conventional microfluidic chip fabrication methods are expensive, require much experience to operate, and are time consuming, while 3D printing offers a solution to these challenges. The 3D printing technique prints models designed using a computer-aided design software such as Autodesk Fusion 360. In this work the authors show example microfluidic chips which were printed using a 3D printer, these include an X-channel chip, Y-channel chip and a lateral flow chip which can all be integrated with biosensing setups.

Smart Microfluidics: Synergy of Machine Learning and Microfluidics in the Development of Medical Diagnostics for Chronic and Emerging Infectious Diseases.

The COVID-19 pandemic, emerging/re-emerging infections as well as other non-communicable chronic diseases, highlight the necessity of smart microfluidic point-of-care diagnostic (POC) devices and systems in developing nations as risk factors for infections, severe disease manifestations and poor clinical outcomes are highly represented in these countries. These POC devices are also becoming vital as analytical procedures executable outside of conventional laboratory settings are seen as the future of healthcare delivery. Microfluidics have grown into a revolutionary system to miniaturize chemical and biological experimentation, including disease detection and diagnosis utilizing μPads/paper-based microfluidic devices, polymer-based microfluidic devices and 3-dimensional printed microfluidic devices. Through the development of droplet digital PCR, single-cell RNA sequencing, and next-generation sequencing, microfluidics in their analogous forms have been the leading contributor to the technical advancements in medicine. Microfluidics and machine-learning-based algorithms complement each other with the possibility of scientific exploration, induced by the framework's robustness, as preliminary studies have documented significant achievements in biomedicine, such as sorting, microencapsulation, and automated detection. Despite these milestones and potential applications, the complexity of microfluidic system design, fabrication, and operation has prevented widespread adoption. As previous studies focused on microfluidic devices that can handle molecular diagnostic procedures, researchers must integrate these components with other microsystem processes like data acquisition, data processing, power supply, fluid control, and sample pretreatment to overcome the barriers to smart microfluidic commercialization.

Microfluidic fabrication of photo-responsive Ansamitocin P-3 loaded liposomes for the treatment of breast cancer.

Ansamitocin P-3 (AP-3) is a promising anticancer agent. However, its low solubility has limited its biomedical applications. The preparation of liposomal formulations for the delivery of low solubility drugs using the microfluidic platform has attracted increasing attention in the pharmaceutical industry. In addition, photodynamic therapy (PDT) is a non-invasive and efficient strategy for the treatment of cancers, making photodynamic liposomes one of the most promising drug delivery systems (DDSs). In this study, a recently developed microfluidic device (swirl mixer) was used for the manufacturing of temperature-sensitive liposomes (TSL) that can be activated by near-infrared stimulation for the treatment of breast cancer. Changing the processing parameters of the microfluidic system allowed for optimizing the properties of the produced liposomes (e.g., particle size and size distribution). For the first time, the anticancer drug AP-3 and the photosensitizer indocyanine green (ICG) were encapsulated into TSL (AP-3/ICG@TSL) during microfluidic processing. The results show that AP-3/ICG@TSL are biocompatible and can significantly reduce the toxicity of AP-3 to normal tissues. After infrared laser irradiation, the heat generated from ICG not only resulted in the cancer cell toxicity, but also facilitated the release of AP-3 in tumor cells. AP-3/ICG@TSL with infrared laser irradiation was found to be able to significantly inhibit the growth of MCF-7 multicellular tumor spheroids (MCTSs) in vitro and MCF-7 tumors subcutaneously inoculated in nude mice as an in vivo model. In addition, it also showed no signs of damage to other organs. The current results demonstrated that the AP-3/ICG@TSL fabricated using the microfluidic swirl mixer is a promising DDS for breast cancer therapy.

Scalable microfluidic fabrication of vertically aligned two-dimensional nanosheets for superior thermal management.

Two-dimensional (2D) nanosheets have been assembled into various macroscopic structures for wide engineering applications. To fully explore their exceptional thermal, mechanical, and electrical properties, 2D nanosheets must be aligned into highly ordered structures due to their strong structural anisotropy. Structures stacked layer by layer such as films and fibers have been readily assembled from 2D nanosheets due to their planar geometry. However, scalable manufacturing of macroscopic structures with vertically aligned 2D nanosheets remains challenging, given their large lateral size with a thickness of only a few nanometers. Herein, we report a scalable and efficient microfluidics-enabled sheet-aligning process to assemble 2D nanosheets into a large-area film with a highly ordered vertical alignment. By applying microchannels with a high aspect ratio, 2D nanosheets were well aligned vertically under strong channel size confinement and high flow shear stress. A vertically aligned graphene sheet film was obtained and applied to effectively improve the heat transfer of thermal interfacial materials (TIMs). Superior through-plane thermal conductivity of 82.7 W m-1 K-1 at a low graphene content of 11.8 vol% was measured for vertically aligned TIMs. Thus, they demonstrate exceptional thermal management performance for switching power supplies with high reliability.

Development of Low-Residual-Stress Photosensitive Adhesive Materials for Wafer-Scale Microfluidic Device Fabrication

We have previously developed a photosensitive adhesive material that enables microfluidic channel structure formation and subsequent bonding to a cover with the aim of simplifying the process of wafer-scale microfluidic device fabrication. However, the bonding process and a subsequent curing process required heat treatment at 200 ℃ and 180 ℃, respectively, resulting in the generation of relatively high residual thermal stress. This may lead to delamination, warping, and cracking depending on the application environment. Therefore, in the present study, we designed a new material platform in which the patterning, bonding, and curing processes are based on ultraviolet (UV) reaction of polyfunctional acrylate, tackiness of epoxy resin, and UV/thermal reaction of epoxy resin in order to achieve low residual stress through low-temperature processing. Proof of concept was conducted through the evaluation of various physical properties of the system and found that the new type of photosensitive adhesive material, which bonded to a cover and cured at lower temperatures of 100 ℃ and 120 ℃, respectively, had a low residual stress of 10 MPa and good insulation reliability. This is a promising material for applications such as cooling devices for semiconductor chips, where long-term reliability is required.

A Customizable and Low-Cost Ultraviolet Exposure System for Photolithography.

For microfluidic device fabrication in the research, industry, and commercial areas, the curing and transfer of patterns on photoresist relies on ultraviolet (UV) light. Often, this step is performed by commercial mask aligner or UV lamp exposure systems; however, these machines are often expensive, large, and inaccessible. To find an alternative solution, we present an inexpensive, customizable, and lightweight UV exposure system that is user-friendly and readily available for a homemade cleanroom. We fabricated a portable UV exposure system that costs under $200. The wafer holder's adjustable height enabled for the selection of the appropriate curing distance, demonstrating our system's ability to be easily tailored for different applications. The high light uniformity across a 4" diameter wafer holder (light intensity error ~2.9%) was achieved by adding a light diffusing film to the apparatus. These values are comparable to the light uniformity across a 5" diameter wafer holder from a commercial mask aligner (ABM 3000HR Mask Aligner), that has a light intensity error of ~4.0%. We demonstrated the ability to perform photolithography with high quality by fabricating microfluidic devices and generating uniform microdroplets. We achieved comparable quality to the wafer patterns, microfluidic devices, and droplets made from the ABM 3000HR Mask Aligner.

Role of Polymers in Microfluidic Devices.

Polymers are sustainable and renewable materials that are in high demand due to their excellent properties. Natural and synthetic polymers with high flexibility, good biocompatibility, good degradation rate, and stiffness are widely used for various applications, such as tissue engineering, drug delivery, and microfluidic chip fabrication. Indeed, recent advances in microfluidic technology allow the fabrication of polymeric matrix to construct microfluidic scaffolds for tissue engineering and to set up a well-controlled microenvironment for manipulating fluids and particles. In this review, polymers as materials for the fabrication of microfluidic chips have been highlighted. Successful models exploiting polymers in microfluidic devices to generate uniform particles as drug vehicles or artificial cells have been also discussed. Additionally, using polymers as bioink for 3D printing or as a matrix to functionalize the sensing surface in microfluidic devices has also been mentioned. The rapid progress made in the combination of polymers and microfluidics presents a low-cost, reproducible, and scalable approach for a promising future in the manufacturing of biomimetic scaffolds for tissue engineering.

Improvement of the signal to noise ratio for fluorescent imaging in microfluidic chips

Microfluidics systems can be fabricated in various ways using original silicon glass systems, with easy Si processing and surface modifications for subsequent applications such as cell seeding and their study. Fluorescent imaging of cells became a standard technique for the investigation of cell behavior. Unfortunately, high sensitivity fluorescent imaging, e.g., using total internal reflection fluorescence (TIRF) microscopy, is problematic in these microfluidic systems because the uneven surfaces of the silicon channels’ bottoms affect light penetration through the optical filters. In this work, we study the nature of the phenomenon, finding that the problem can be rectified by using a silicon-on-insulator (SOI) substrate, defining the channel depth by the thickness of the top Si layer, and halting the etching at the buried SiO2 layer. Then the fluorescent background signal drops by = 5 times, corresponding to the limit of detection drop from = 0.05 mM to = 50 nM of fluorescein. We demonstrate the importance of a flat surface using TIRF-based single-molecule detection, improving the signal to a noise ratio more than 18 times compared to a conventional Si wafer. Overall, using very high-quality SOI substrates pays off, as it improves the fluorescence image quality due to the increase in signal-to-noise ratio. Concerning the cost of microfluidic device fabrication—design, mask fabrication, wafer processing, and device testing—the initial SOI wafer cost is marginal, and using it improves the system performance.

(Invited) Surface Modification to Control Wetting and Adhesion of Aqueous Solutions

rf plasmas and electrochemical treatments have been used extensively for thin film deposition and etching in the fabrication of microelectronic and photonic devices. These methods can be applied to non-traditional substrates (e.g., paper and stainless steel) to alter surface and bulk properties to create (i) paper-based analytical devices with application to microfluidic devices, and (ii) antibacterial structures on stainless steel. Two examples will be described. First, the effects of plasma-based oxygen etching of cellulose surfaces followed by plasma-enhanced fluorocarbon film deposition will be described and shown to control wetting and adhesion of water droplets. These process steps can also be used to form fully enclosed hydrophilic channels in bulk paper and thereby allow microfluidic device fabrication on inexpensive and renewable substrates. Electrochemical etching of stainless steel creates micro- and nano-structures on stainless steel surfaces that repel water and bacteria. These surfaces have potential applications in orthopedic implants and consumer metal products prone to bacterial contamination.

Macromolecular microfluidic concentrators

Microfluidic design, fabrication, and experiments have developed rapidly, leading to lab-on-chip separation or fractionation. In this work, we design a continuous concentrator for macromolecular solutions. Our design relies on the analytical solutions for orientational diffusion under laminar pressure-driven slot flow through a microchannel [W. Stasiak and C. Cohen, “Dilute solutions of macromolecules in a rectilinear Poiseuille flow,” J. Chem. Phys. 78, 553 (1983)]. Using rigid dumbbell theory, we provide analytical solutions for the design of our microfluidic macromolecular hydrodynamic chromatography. We arrive at our design through the use of well-known confinement-driven composition profiles. Using a pair of razor-sharp blades, our design separates the slot flow into a symmetric core inner slot (concentrated) between two outer slots (diluted). We discover a minimum dimensionless blade leading edge separation for complete fractionation, and that this decreases with confinement and also decreases with dimensionless shear rate.

One Step Fabrication of Hollow and Highly Flexible Polydimethylsiloxane Microneedles

In this study, the hollow and highly flexible polydimethylsiloxane microneedles were fabricated in a one step and simple design. For this purpose, a commercial dermastamping device (Dermastamp® 140 DRS) was used as a mold to obtain highly flexible PDMS microneedles containing channels. With the proposed design, microneedles with a total height of 1500 μm, 1500 μm center-to-center spacing and 150 μm channel diameter was successfully fabricated. These data are all compatible with the dimensions and the geometry of the mold used. Then, a syringe adapter was fabricated with a 3D printer and combined with the hollow PDMS microneedle patch for the high-throughput production of alginate beads. After the adapter and the hollow PDMS microneedle patch combination was placed into the syringe pump, the mostly spherical alginate beads with a mean diameter of 2.0 ± 0.3 mm was produced. To sum up, the proposed design and fabrication scheme first offer a novel and simple strategy for the fabrication of hollow polymeric microneedles. Moreover, this system has the potential to be used not only for the high-throughput microfluidic fabrication of polymeric beads, but also in controlled drug delivery systems and cell encapsulation studies.

PDMS Organ-On-Chip Design and Fabrication: Strategies for Improving Fluidic Integration and Chip Robustness of Rapidly Prototyped Microfluidic In Vitro Models.

The PDMS-based microfluidic organ-on-chip platform represents an exciting paradigm that has enjoyed a rapid rise in popularity and adoption. A particularly promising element of this platform is its amenability to rapid manufacturing strategies, which can enable quick adaptations through iterative prototyping. These strategies, however, come with challenges; fluid flow, for example, a core principle of organs-on-chip and the physiology they aim to model, necessitates robust, leak-free channels for potentially long (multi-week) culture durations. In this report, we describe microfluidic chip fabrication methods and strategies that are aimed at overcoming these difficulties; we employ a subset of these strategies to a blood–brain-barrier-on-chip, with others applied to a small-airway-on-chip. Design approaches are detailed with considerations presented for readers. Results pertaining to fabrication parameters we aimed to improve (e.g., the thickness uniformity of molded PDMS), as well as illustrative results pertaining to the establishment of cell cultures using these methods will also be presented.

The Fabrication and Bonding of Thermoplastic Microfluidics: A Review.

Various fields within biomedical engineering have been afforded rapid scientific advancement through the incorporation of microfluidics. As literature surrounding biological systems become more comprehensive and many microfluidic platforms show potential for commercialization, the development of representative fluidic systems has become more intricate. This has brought increased scrutiny of the material properties of microfluidic substrates. Thermoplastics have been highlighted as a promising material, given their material adaptability and commercial compatibility. This review provides a comprehensive discussion surrounding recent developments pertaining to thermoplastic microfluidic device fabrication. Existing and emerging approaches related to both microchannel fabrication and device assembly are highlighted, with consideration toward how specific approaches induce physical and/or chemical properties that are optimally suited for relevant real-world applications.

Nanofiber self-consistent additive manufacturing process for 3D microfluidics

3D microfluidic devices have emerged as powerful platforms for analytical chemistry, biomedical sensors, and microscale fluid manipulation. 3D printing technology, owing to its structural fabrication flexibility, has drawn extensive attention in the field of 3D microfluidics fabrication. However, the collapse of suspended structures and residues of sacrificial materials greatly restrict the application of this technology, especially for extremely narrow channel fabrication. In this paper, a 3D printing strategy named nanofiber self-consistent additive manufacturing (NSCAM) is proposed for integrated 3D microfluidic chip fabrication with porous nanofibers as supporting structures, which avoids the sacrificial layer release process. In the NSCAM process, electrospinning and electrohydrodynamic jet (E-jet) writing are alternately employed. The porous polyimide nanofiber mats formed by electrospinning are ingeniously applied as both supporting structures for the suspended layer and percolating media for liquid flow, while the polydimethylsiloxane E-jet writing ink printed on the nanofiber mats (named construction fluid in this paper) controllably permeates through the porous mats. After curing, the resultant construction fluid–nanofiber composites are formed as 3D channel walls. As a proof of concept, a microfluidic pressure-gain valve, which contains typical features of narrow channels and movable membranes, was fabricated, and the printed valve was totally closed under a control pressure of 45 kPa with a fast dynamic response of 52.6 ms, indicating the feasibility of NSCAM. Therefore, we believe NSCAM is a promising technique for manufacturing microdevices that include movable membrane cavities, pillar cavities, and porous scaffolds, showing broad applications in 3D microfluidics, soft robot drivers or sensors, and organ-on-a-chip systems.

Microfluidic Formulation of Topological Hydrogels for Microtissue Engineering.

Microfluidics has recently emerged as a powerful tool in generation of submillimeter-sized cell aggregates capable of performing tissue-specific functions, so-called microtissues, for applications in drug testing, regenerative medicine, and cell therapies. In this work, we review the most recent advances in the field, with particular focus on the formulation of cell-encapsulating microgels of small "dimensionalities": "0D" (particles), "1D" (fibers), "2D" (sheets), etc., and with nontrivial internal topologies, typically consisting of multiple compartments loaded with different types of cells and/or biopolymers. Such structures, which we refer to as topological hydrogels or topological microgels (examples including core-shell or Janus microbeads and microfibers, hollow or porous microstructures, or granular hydrogels) can be precisely tailored with high reproducibility and throughput by using microfluidics and used to provide controlled "initial conditions" for cell proliferation and maturation into functional tissue-like microstructures. Microfluidic methods of formulation of topological biomaterials have enabled significant progress in engineering of miniature tissues and organs, such as pancreas, liver, muscle, bone, heart, neural tissue, or vasculature, as well as in fabrication of tailored microenvironments for stem-cell expansion and differentiation, or in cancer modeling, including generation of vascularized tumors for personalized drug testing. We review the available microfluidic fabrication methods by exploiting various cross-linking mechanisms and various routes toward compartmentalization and critically discuss the available tissue-specific applications. Finally, we list the remaining challenges such as simplification of the microfluidic workflow for its widespread use in biomedical research, bench-to-bedside transition including production upscaling, further in vivo validation, generation of more precise organ-like models, as well as incorporation of induced pluripotent stem cells as a step toward clinical applications.

Bonding Strategies for Thermoplastics Applicable for Bioanalysis and Diagnostics.

Microfluidics is a multidisciplinary science that includes physics, chemistry, engineering, and biotechnology. Such microscale systems are receiving growing interest in applications such as analysis, diagnostics, and biomedical research. Thermoplastic polymers have emerged as one of the most attractive materials for microfluidic device fabrication owing to advantages such as being optically transparent, biocompatible, cost-effective, and mass producible. However, thermoplastic bonding is a key challenge for sealing microfluidic devices. Given the wide range of bonding methods, the appropriate bonding approach should be carefully selected depending on the thermoplastic material and functional requirements. In this review, we aim to provide a comprehensive overview of thermoplastic fabricating and bonding approaches, presenting their advantages and disadvantages, to assist in finding suitable microfluidic device bonding methods. In addition, we highlight current applications of thermoplastic microfluidics to analyses and diagnostics and introduce future perspectives on thermoplastic bonding strategies.

A Protocol for Fabrication and on-Chip Cell Culture to Recreate PAH-Afflicted Pulmonary Artery on a Microfluidic Device.

Pulmonary arterial hypertension (PAH) is a rare pulmonary vascular disease that affects people of all ethnic origins and age groups including newborns. In PAH, pulmonary arteries and arterioles undergo a series of pathological changes including remodeling of the entire pulmonary vasculatures and extracellular matrices, mis-localized growth of pulmonary arterial cells, and development of glomeruloid-like lesions called plexiform lesions. Traditionally, various animal and cellular models have been used to understand PAH pathophysiology, investigate sex-disparity in PAH and monitor therapeutic efficacy of PAH medications. However, traditional models can only partially capture various pathological features of PAH, and they are not adaptable to combinatorial study design for deciphering intricately intertwined complex cellular processes implicated in PAH pathogenesis. While many microfluidic chip-based models are currently available for major diseases, no such disease-on-a-device model is available for PAH, an under investigated disease. In the absence of any chip-based models of PAH, we recently proposed a five-channel polydimethylsiloxane (PDMS)-based microfluidic device that can emulate major pathological features of PAH. However, our proposed model can make a bigger impact on the PAH field only when the larger scientific community engaged in PAH research can fabricate the device and develop the model in their laboratory settings. With this goal in mind, in this study, we have described the detailed methodologies for fabrication and development of the PAH chip model including a thorough explanation of scientific principles for various steps for chip fabrication, a detailed list of reagents, tools and equipment along with their source and catalogue numbers, description of laboratory setup, and cautionary notes. Finally, we explained the methodologies for on-chip cell seeding and application of this model for studying PAH pathophysiology. We believe investigators with little or no training in microfluidic chip fabrication can fabricate this eminently novel PAH-on-a-chip model. As such, this study will have a far-reaching impact on understanding PAH pathophysiology, unravelling the biological mystery associated with sexual dimorphism in PAH, and developing PAH therapy based on patient sex and age.

Multi-Resin Masked Stereolithography (MSLA) 3D Printing for Rapid and Inexpensive Prototyping of Microfluidic Chips with Integrated Functional Components.

Stereolithography based 3D printing of microfluidics for prototyping has gained a lot of attention due to several advantages such as fast production, cost-effectiveness, and versatility over traditional photolithography-based microfabrication techniques. However, existing consumer focused SLA 3D printers struggle to fabricate functional microfluidic devices due to several challenges associated with micron-scale 3D printing. Here, we explore the origins and mechanism of the associated failure modes followed by presenting guidelines to overcome these challenges. The prescribed method works completely with existing consumer class inexpensive SLA printers without any modifications to reliably print PDMS cast microfluidic channels with channel sizes as low as ~75 μm and embedded channels with channel sizes as low ~200 μm. We developed a custom multi-resin formulation by incorporating Polyethylene glycol diacrylate (PEGDA) and Ethylene glycol polyether acrylate (EGPEA) as the monomer units to achieve micron sized printed features with tunable mechanical and optical properties. By incorporating multiple resins with different mechanical properties, we were able to achieve spatial control over the stiffness of the cured resin enabling us to incorporate both flexible and rigid components within a single 3D printed microfluidic chip. We demonstrate the utility of this technique by 3D printing an integrated pressure-actuated pneumatic valve (with flexible cured resin) in an otherwise rigid and clear microfluidic device that can be fabricated in a one-step process from a single CAD file. We also demonstrate the utility of this technique by integrating a fully functional finger-actuated microfluidic pump. The versatility and accessibility of the demonstrated fabrication method have the potential to reduce our reliance on expensive and time-consuming photolithographic techniques for microfluidic chip fabrication and thus drastically lowering our barrier to entry in microfluidics research.

Rapid, inexpensive fabrication of electrophoretic microdevices for fluorescence detection.

The laser print, cut, and laminate (PCL) method for microfluidic device fabrication can be leveraged for rapid and inexpensive prototyping of electrophoretic microchips useful for optimizing separation conditions. The rapid prototyping capability allows the evaluation of fluidic architecture, applied fields, reagent concentrations, and sieving matrix, all within the context of using fluorescence‐compatible substrates. Cyclic olefin copolymer and toner‐coated polyethylene terephthalate (tPeT) were utilized with the PCL technique and bonding methods optimized to improve device durability during electrophoresis. A series of separation channel designs and centrifugation conditions that provided successful loading of sieving polymer in less than 3 min was described. Separation of a 400‐base DNA sizing ladder provided calculated base resolution between 3 and 4 bases, a greater than 18‐fold improvement over separations on similar substrates. Finally, the accuracy and precision capabilities of these devices were demonstrated by separating and sizing DNA fragments of 147 and 167 bases as 148.62 ± 2 and 166.48 ± 3 bases, respectively.

Dual-wavelength volumetric stereolithography of multilevel microfluidic devices.

Microfluidic devices are typically fabricated in an expensive, multistep process (e.g., photolithography, etching, and bonding). Additive manufacturing (AM) has emerged as a revolutionary technology for simple and inexpensive fabrication of monolithic structures-enabling microfluidic designs that are challenging, if not impossible, to make with existing fabrication techniques. Here, we introduce volumetric stereolithography (vSLA), an AM method in which polymerization is constrained to specific heights within a resin vat, allowing layer-by-layer fabrication without a moving platform. vSLA uses an existing dual-wavelength chemistry that polymerizes under blue light (λ = 458 nm) and inhibits polymerization under UV light (λ = 365 nm). We apply vSLA to fabricate microfluidic channels with different spatial and vertical geometries in less than 10 min. Channel heights ranged from 400 μm to 1 mm and could be controlled with an optical dose, which is a function of blue and UV light intensities and exposure time. Oxygen in the resin was found to significantly increase the amount of dose required for curing (i.e., polymerization to a gelled state), and we recommend that an inert vSLA system is used for rapid and reproducible microfluidic fabrication. Furthermore, we recommend polymerizing far beyond the gel point to form more rigid structures that are less susceptible to damage during post-processing, which can be done by simultaneously increasing the blue and UV light absorbance of the resin with light intensities. We believe that vSLA can simplify the fabrication of complex multilevel microfluidic devices, extending microfluidic innovation and availability to a broader community.