Microfluidic Design Research Articles

Coupling of Solid-Phase Microextraction Directly to Mass Spectrometry via an Improved Microfluidic Open Interface to Facilitate High-Throughput Determinations.

Mass spectrometry analysis can be performed by introducing samples directly to mass spectrometry, allowing the increase of the analysis throughput; however, some disadvantages of direct-to-mass spectrometry analysis include susceptibility to matrix effects and risk of instrument contamination from inadequate sample preparation. Solid-phase microextraction is one of the most suitable sample preparation methods for direct-to-mass spectrometry analysis, as it offers matrix-compatible coatings which ensure analyte enrichment with minimal or no interference from matrix. One of the ways solid-phase microextraction can be coupled directly to mass spectrometry is via a microfluidic open interface. This manuscript reports improvements made to the initial microfluidic open interface design, where the system components have been simplified to mostly commercially available materials. In addition, the analysis of samples has been automated by implementing software that fully controls the analysis workflow, where the washing procedure is optimized to completely reduce the carryover. Herein, the extraction and desorption time profiles from thin and thick SPME devices was studied where the overall workflow consisted of high-throughput sample preparation of 1.3 min per 96 samples and <1 min per sample instrumental analysis.

A multistep immunoassay on a 3D-printed capillarity-driven microfluidic device for point-of-care diagnostics

Although efficient diagnostic testing often relies on centralized laboratories, rapid diagnostics at the point-of-care (POC) are increasingly investigated to provide an affordable and easy-to-use alternative that requires minimal external equipment. Implementing complex assays on such POC devices is challenging, and existing methods such as paper-based analytical devices suffer from laborious and heterogeneous fabrication. 3D printing is a promising technology to enable the single-step fabrication of such diagnostic devices. We previously reported monolithic, 3D-printed microfluidics driven by capillarity, and used the platform to implement a proof-of-concept enzyme-linked immunosorbent assay (ELISA) for immunoglobulin E (IgE) detection. In this study, we aimed to improve the time-to-result and detection limit of this approach by (i) designing a second-generation 3D-printed device and (ii) further optimizing the IgE ELISA. Herein, we selected horseradish peroxidase for faster and more sensitive signal amplification compared to the first-generation device, for which we adapted the microfluidic device design to allow sequential flow while using separated reagent channels for each assay step. To implement the ELISA on this new design, we decreased reagent volumes and incubation times to be compatible with POC, and then we translated this assay stepwise from microtiter plates to the completely 3D-printed, second-generation device. Overall, this work gives additional insight into the biocompatibility of these 3D-printed devices, as well as demonstrates the huge potential of powder bed 3D printing to fabricate POC diagnostic devices.

Self-synchronization of reinjected droplets for high-efficiency droplet pairing and merging

Droplet merging serves as a powerful tool to add reagents to moving droplets for biological and chemical reactions. However, unsynchronized droplet pairing impedes high-efficiency merging. Here, we develop a microfluidic design for the self-synchronization of reinjected droplets. A periodic increase in the hydrodynamic resistance caused by droplet blocking a T-junction enables automatic pairing of droplets. After inducing spacing, the paired droplets merge downstream under an electric field. The blockage-based design can achieve a 100% synchronization efficiency even when the mismatch rate of droplet frequencies reaches 10%. Over 98% of the droplets can still be synchronized at nonuniform droplet sizes and fluctuating reinjection flow rates. Moreover, the droplet pairing ratio can be adjusted flexibly for on-demand sample addition. Using this system, we merge two groups of droplets encapsulating enzyme/substrate, demonstrating its capacity to conduct multi-step reactions. We also combine droplet sorting and merging to coencapsulate single cells and single beads, providing a basis for high-efficiency single-cell sequencing. We expect that this system can be integrated with other droplet manipulation systems for a broad range of chemical and biological applications.

Chemical Imaging of Mass Transport Near the No-Slip Interface of a Microfluidic Device using Attenuated Total Reflection–Fourier Transform Infrared Spectroscopy

Mass transport in geometrically confined environments is fundamental to microfluidic applications. Measuring the distribution of chemical species on flow requires the use of spatially resolved analytical tools compatible with microfluidic materials and designs. Here, the implementation of an attenuated total reflection-Fourier transform infrared spectroscopy (ATR-FTIR) imaging (macro-ATR) approach for chemical mapping of species in microfluidic devices is described. The imaging method is configurable between a large field of view, single-frame imaging, and the use of image stitching to build composite chemical maps. Macro-ATR is used to quantify transverse diffusion in the laminar streams of coflowing fluids in dedicated microfluidic test devices. It is demonstrated that the ATR evanescent wave, which primarily probes the fluid within ∼500 nm of the channel surface, provides accurate quantification of the spatial distribution of species in the entire microfluidic device cross section. This is the case when flow and channel conditions promote vertical concentration contours in the channel as verified by three-dimensional numeric simulations of mass transport. Furthermore, the validity of treating the mass transport problem in a simplified and faster approach using reduced dimensionality numeric simulations is described. Simplified one-dimensional simulations, for the specific parameters used herein, overestimate diffusion coefficients by a factor of approximately 2, whereas full three-dimensional simulations accurately agree with experimental results.

Naturally derived colloidal rods in microfluidic flows.

Naturally derived colloidal rods (CR) are promising building blocks for developing sustainable soft materials. Engineering new materials based on naturally derived CR requires an in-depth understanding of the structural dynamics and self-assembly of CR in dispersion under processing conditions. With the advancement of microfabrication techniques, many microfluidic platforms have been employed to study the structural dynamics of CR under flow. However, each microfluidic design has its pros and cons which need careful evaluation in order to fully meet the experimental goal and correctly interpret the data. We analyze recent results obtained from naturally derived CR and relevant rod-like macromolecules under microfluidic flows, with emphasis on the dynamical behavior in shear- and extensional-dominated flows. We highlight the key concepts required in order to assess and evaluate the results obtained from different CR and microfluidic platforms as a whole and to aid interconnections with neighboring fields. Finally, we identify and discuss areas of interest for future research directions.

Experimental Investigation of Microfluidic Device for Platelet Activation.

Wound healing is accelerated when Platelet Rich Plasma is activated and growth factors are released. In this study, it was aimed to stimulate platelets without using chemical stimulants. Two types of mechanical platelet activation methods have been proposed in this study. The first one is a microfluidic chip developed with the shear-induced platelet activation approach. The second one is a piezo-based ultrasound-assisted device which provides platelet activation by stimulating with an ultrasonic wave (0.55 and 1.1 MHz). Three different microfluidic chip designs were worked out to determine the optimal shear stress characteristics; 8-nodes (2789 µs, 288 shear pulses, and 98.3 dyne/cm2), 40-nodes (2765 µs, 1440 shear pulses, and 95.5 dyne/cm2) and pillar-shaped (1030 µs, 1656 shear pulses, and 48.1 dyne/cm2). The highest platelet activation rate (72.7%) was obtained from the chips with 8-nodes. In the ultrasound-assisted device, 32.4% activation rate was obtained from ultrasound waves with 0.55 MHz frequency and 10 Vp-p amplitude. These activation rates, determined by CD62P (P-Selectin) expression, are significantly higher than spontaneous activation of intact platelets (8.5%). In addition, the gradual increase in activation of stimulated platelets with incubation at room temperature showed that activation continued after stimulation. The results showed that these microfluidic devices can be used for platelet activation to enhance the effect of PRP treatment and might reduce adverse immune reactions that may happened due to the use of exogenous activator substances. Fast-response, low-cost, easy-to-use and controllable biomedical device have been developed for PRP applications.

Injectable engineered micro/nano-complexes trigger the reprogramming of bone immune epigenetics

Imbalance in the local immune microenvironment plays a vital role in the nonunion or delayed healing of bone defects. Additionally, there exists a cross-connection between material design of microfluidics and immune regulation. Therefore, we aimed to investigate whether epigenetic reprogramming of the microfluidic microsphere systems could regulate microenvironmental homeostasis in the acute phase of fracture and facilitate bone regeneration. To this end, we used Gelma hydrogel microspheres as the base carriers to design and fabricate an injectable, macrophage-targeted, engineered micro/nano microsphere reprogramming system (Gelma@Lip@Pla). Liposome (Lip) coating and macrophage targeting were improved by adding DSPE-PEG-CHO and phosphatidylserine (Pla). the surface of Gelma was coated with a lipid membrane, Schiff base, and hydrogel matrix network; the resultant macrophage-targeted liposomes activated AKT signaling in the local microenvironment by releasing the AKT signal activator dihydrocapsaicin and amplifiers (pcDNA3.1-AKT), triggering gene reprogramming in the local microenvironment and restarting the bone healing process. Compared with pcDNA3.1-AKT and dihydrocapsaicin-loaded liposomes (Lip@Pla) and pure Gelma hydrogel microgels (Gelma), the Gelma@Lip@Pla group significantly prolonged the release time of active ingredients. They also promoted the polarization of macrophages to the M2 phenotype, leading to the reactivation of bone marrow-derived mesenchymal stem cell osteogenic differentiation and human umbilical vein endothelial cell angiogenesis. Furthermore, in vivo experiments confirmed that local injection of Gelma@Lip@Pla could significantly accelerate the regeneration of a defective rat femoral condyle. Collectively, the Gelma@Lip@Pla microgel exhibits considerable potential as an extended delivery platform for treating bone defects and other immune-related diseases involving macrophages.

Increasing the Capture Rate of Circulating Tumor DNA in Unaltered Plasma Using Passive Microfluidic Mixer Flow Cells.

A limiting factor in using blood-based liquid biopsies for cancer detection is the volume of extracted blood required to capture a measurable number of circulating tumor DNA (ctDNA). To overcome this limitation, we developed a technology named the dCas9 capture system to capture ctDNA from unaltered flowing plasma, removing the need to extract the plasma from the body. This technology has provided the first opportunity to investigate whether microfluidic flow cell design can affect the capture of ctDNA in unaltered plasma. With inspiration from microfluidic mixer flow cells designed to capture circulating tumor cells and exosomes, we constructed four microfluidic mixer flow cells. Next, we investigated the effects of these flow cell designs and the flow rate on the rate of captured spiked-in BRAF T1799A (BRAFMut) ctDNA in unaltered flowing plasma using surface-immobilized dCas9. Once the optimal mass transfer rate of ctDNA, identified by the optimal ctDNA capture rate, was determined, we investigated whether the design of the microfluidic device, flow rate, flow time, and the number of spiked-in mutant DNA copies affected the rate of capture by the dCas9 capture system. We found that size modifications to the flow channel had no effect on the flow rate required to achieve the optimal capture rate of ctDNA. However, decreasing the size of the capture chamber decreased the flow rate required to achieve the optimal capture rate. Finally, we showed that, at the optimal capture rate, different microfluidic designs using different flow rates could capture DNA copies at a similar rate over time. In this study, the optimal capture rate of ctDNA in unaltered plasma was identified by adjusting the flow rate in each of the passive microfluidic mixer flow cells. However, further validation and optimization of the dCas9 capture system are required before it is ready to be used clinically.

Liquid Shuttle Mediated by Microwick for Open‐Air Microfluidics

AbstractMicrofluidics has experienced rapid progress in additive manufacturing and microfluidic soft robots. The design of microfluidics is already moving into a more intelligent, integrated, and detachable direction. However, the pipeline resistance needs more external energy input to achieve high flow speed. Guided transport of liquid in the open‐air‐space microfluidics will be an effective solution. Inspired by the water shuttle on the pitcher plant tendril, herein, an open‐air microfluidic transport device is designed that consists of a superhydrophilic microwick with multi‐microgrooves by stereolithography. The liquid film confined in microgrooves can promote rapid fluid shuttle on the wet surface to enhance transport rate and inhibit the Rayleigh‐Plateau instability from forming larger dripping drops. The dripping volume and threshold Capillary number are optimized for effective liquid transport and drainage. State‐of‐the‐art microwick liquid shuttle technologies can guide liquid continuously in a prescribed direction or into multiple directions with 98% transport efficiency (the ratio of liquid collection volume and liquid injection volume) for water and 97% for ethanol in the closed‐open‐closed space. The proposed mechanism has the potential to streamline microfluidic applications—and, therefore, accelerate relevant liquid delivery development and ultimately their applications in microfluidic chip and additive manufacturing.

Channel Routing for Microfluidic Devices: A Comprehensive and Accessible Design Tool

Microfluidics is a technology that enables moving analytic processes from expensive and bulky laboratory equipment to small-scale devices. Microfluidic devices, usually in the form of labs-on-a-chips (LoCs), have found many great applications in medicine, biology, and chemistry. In particular, LoCs that utilize channels to transport fluids or droplets between different components on the chip are a promising technology. However, the design process of such channel-based LoCs is in need of further automation efforts since the underlying design steps are still rather complex and conducted mainly by hand. An important task in microfluidic design automation is the so-called channel routing, where components on LoCs are connected by microfluidic channels. Methods that aim to automate this routing task must factor in the specific demands of microfluidic devices. Common requirements for microfluidic routing layouts are to prevent sharp channel bends and to realize a particular length of channels. Unfortunately, most of the available routing algorithms address these requirements only partly and insufficiently. In this work, we propose a router that is able to overcome these shortcomings and allows automatic channel routing with a minimal bending radius as well as a desired length. In order to make the router accessible to users with little to no design automation expertise, the solution is implemented as an online tool with a user-friendly and intuitive interface. The resulting tool can be accessed at <uri xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">https://www.cda.cit.tum.de/research/microfluidics/channel_router/</uri> .

Microfluidics for COVID-19: From Current Work to Future Perspective

Spread of coronavirus disease 2019 (COVID-19) has significantly impacted the public health and economic sectors. It is urgently necessary to develop rapid, convenient, and cost-effective point-of-care testing (POCT) technologies for the early diagnosis and control of the plague's transmission. Developing POCT methods and related devices is critical for achieving point-of-care diagnosis. With the advantages of miniaturization, high throughput, small sample requirements, and low actual consumption, microfluidics is an essential technology for the development of POCT devices. In this review, according to the different driving forces of the fluid, we introduce the common POCT devices based on microfluidic technology on the market, including paper-based microfluidic, centrifugal microfluidic, optical fluid, and digital microfluidic platforms. Furthermore, various microfluidic-based assays for diagnosing COVID-19 are summarized, including immunoassays, such as ELISA, and molecular assays, such as PCR. Finally, the challenges of and future perspectives on microfluidic device design and development are presented. The ultimate goals of this paper are to provide new insights and directions for the development of microfluidic diagnostics while expecting to contribute to the control of COVID-19.

Chip for dielectrophoretic microbial capture, separation and detection I: theoretical basis of electrode design

We model the dielectrophoretic response of E. coli bacterial cells and red blood cells, upon exposure to an electric field. We model the separation, capture, and release mechanisms under flow conditions in a microfluidic channel and show under which conditions efficient separation of different cell types occurs. The modelling work is aimed to guide the separation electrode architecture and design for experimental validation of the model. The dielectrophoretic force is affected both by the geometry of the electrodes (the gradient of the electric field), the Re{CM(ω)} factor, and the permittivity of the medium ϵm. Our modelling makes testable predictions and shows that designing the electrode structure to ensure structure periodicity with spacing between consecutive traps smaller than the length of the depletion zone ensures efficient capture and separation. Such electrode system has higher capture and separation efficiency than systems with the established circular electrode architecture. The simulated, modelled microfluidic design allows for the separated bacteria, concentrated by dedicated dielectrophoretic regions, to be subsequently detected using label-free functionalized nanowire sensors. The experimental validation of the modelling work presented here and the validation of the theoretical design constraints of the chip electrode architecture is presented in the companion paper in the same issue (Weber MU et al 2022 Chip for dielectrophoretic Microbial Capture, Separation and Detection II: Experimental Study).

Nucleation control of quantum dot synthesis in a microfluidic continuous flow reactor

The use of microfluidics in chemical synthesis is topical due to the potential to improve reproducibility and the ability promptly interrogate a wide range of reaction parameters, the latter of which is necessary for the training of artificial intelligence (AI) algorithms. Applying microfluidic techniques to semiconductor nanocrystals, or quantum dots (QDs), is challenging due to the need for a high-temperature nucleation event followed by particle growth at lower temperatures. Such a high-temperature gradient can be realized using complex, segmented microfluidic reactor designs, which represents an engineering approach. Here, an alternative chemical approach is demonstrated using the cluster seed method of nanoparticle synthesis in a simple microfluidic reactor system. This enables quantum dot nucleation at lower temperatures due to the presence of molecular organometallic compounds (NMe4)4[Cd10Se4(SPh)16] and (NMe4)4[Zn10Se4(SPh)16]. This integration of cluster seeding with microfluidics affords a new mechanism to tailor the reaction conditions for optimizing yields and tuning product properties.

FLUIDOT: A Modular Microfluidic Platform for Single-Cell Study and Retrieval, with Applications in Drug Tolerance Screening and Antibody Mining.

Advancements in lab-on-a-chip technologies have revolutionized the single-cell analysis field. However, an accessible platform for in-depth screening and specific retrieval of single cells, which moreover enables studying diverse cell types and performing various downstream analyses, is still lacking. As a solution, FLUIDOT is introduced, a versatile microfluidic platform incorporating customizable microwells, optical tweezers and an interchangeable cell-retrieval system. Thanks to its smart microfluidic design, FLUIDOT is straightforward to fabricate and operate, rendering the technology widely accessible. The performance of FLUIDOT is validated and its versatility is subsequently demonstrated in two applications. First, drug tolerance in yeast cells is studied, resulting in the discovery of two treatment-tolerant populations. Second, B cells from convalescent COVID-19 patients are screened, leading to the discovery of highly affine, in vitro neutralizing monoclonal antibodies against SARS-CoV-2. Owing to its performance, flexibility, and accessibility, it is foreseen that FLUIDOT will enable phenotypic and genotypic analysis of diverse cell samples and thus elucidate unexplored biological questions.

Microfluidics in the eye: a review of glaucoma implants from an engineering perspective.

Glaucoma is a progressive optic neuropathy in the eye, which is a leading cause of irreversible blindness worldwide and currently affects over 70 million individuals. Clinically, intraocular pressure (IOP) reduction is the only proven treatment to halt the progression of glaucoma. Microfluidic devices such as glaucoma drainage devices (GDDs) and minimally invasive glaucoma surgery (MIGS) devices are routinely used by ophthalmologists to manage elevated IOP, by creating an artificial pathway for the over-accumulated aqueous humor (AH) in a glaucomatous eye, when the natural pathways are severely blocked. Herein, a detailed modelling and analysis of both the natural microfluidic pathways of the AH in the eye and artificial microfluidic pathways formed additionally by the various glaucoma implants are conducted to provide an insight into the causes of the IOP abnormality and the improvement schemes of current implant designs. The mechanisms of representative glaucoma implants have been critically reviewed from the perspective of microfluidics, and we have categorized the current implants into four groups according to the targeted drainage sites of the AH, namely Schlemm's canal, suprachoroidal space, subconjunctival space, and ocular surface. In addition, we propose to divide the development and evolution of glaucoma implant designs into three technological waves, which include microtube (1st), microvalve (2nd) and microsystem (3rd). With the emerging trends of minimal invasiveness and artificial intelligence in the development of medical implants, we envision that a comprehensive glaucoma treatment microsystem is on the horizon, which is featured with active and wireless control of IOP, real-time continuous monitoring of IOP and aqueous rate, etc. The current review could potentially cast light on the unmatched needs, challenges, and future directions of the microfluidic structural and functional designs of glaucoma implants, which would enable an enhanced safety profile, reduced complications, increased efficacy of lowering IOP and reduced IOP fluctuations, closed-loop and on-demand control of IOP, etc.

A facile microfluidic chip design for DNA detection using dengue serotypes as a proof-of-concept case study

Dengue fever is caused by any of the four serotypes of dengue viruses, DENV-1, DENV-2, DENV-3 and DENV-4 spread by mosquito bites and is important to distinguish between them due to lack of cross-protective neutralizing antibodies for each serotype. Secondary infections also put individuals at higher risk for severe dengue illness than those who have not been previously infected. Current preferred assays include reverse transcription-PCR (RT-PCR) and ELISA. To enable on-field diagnosis of dengue serotypes, the detection process would need to be simplified or at least semi-automated. A downstream detection module was conceptualized and fabricated to detect the amplified DNA from the provided PCR mix (product) of previously developed modular microfluidic chips involving sample loading, cell lysis, RNA extraction and RT-PCR. Further, to ensure accuracy, each serotype assay necessitates a positive control, negative control and test sample, which constitutes 3 separate channels for the diagnosis of just 1 serotype. In this study, a 6-channel bi-assay microfluidic chip was designed with pre-loaded diluent and cyanine dye, sample chamber for loading, sequential fluidic sample mixing, and integrated membranes for simultaneous (6-channel) fluidic manipulation from a single actuation source. Positive samples will turn the dye from blue to violet while the negative controls will remain blue. The integrated membranes provided color contrast and facilitated the manipulation of the samples to the same line of sight for simultaneous analysis, paving the way for automated color analysis via smartphone.

Construction of a cleavable linker chemistry-based HBEXO-Chip to isolate circulating exosomes for breast cancer diagnosis.

Breast cancer is presently the most common form of malignant tumour globally, and its precise diagnosis is vital for enhancing patient survival rates and their quality of life. Exosomes, which are small extracellular vesicles containing proteins and nucleic acid molecules, have emerged as ideal cancer markers for liquid biopsy-based diagnostics. Nevertheless, the current methods for isolating exosomes present challenges for clinical implementation. Although immunoaffinity-based microfluidics hold potential for exosome-based cancer diagnostics, existing microfluidic chips struggle to capture and release intact, high-purity, and highly specific exosomes effectively. To surmount these obstacles, we developed the HBEXO-Chip, an innovative immunoaffinity microfluidic device that employs cleavable linker chemistry technology. This chip enables rapid isolation and detection of breast cancer-derived exosomes in peripheral blood. The fishbone-like microfluidic chip design of the HBEXO-Chip heightens the binding likelihood between specific exosomes and antibodies, significantly augmenting capture efficiency. Furthermore, the gentle reaction conditions of the cleavable linker chemistry retain the exosomes' membrane structure's integrity during the release process, which is advantageous for downstream experimental analysis. Our study demonstrated the effectiveness of the HBEXO-Chip in distinguishing breast cancer patients, patients with benign breast tumours, and healthy controls. By quantitatively analysing Epcam+ exosomes in clinical plasma samples, this technology platform provides a quick, user-friendly, highly sensitive, and specific assay for detecting tumour exosomes in peripheral blood, making it a valuable liquid biopsy tool for clinicians to diagnose breast cancer.

Behavioural responses of zebrafish with sound stimuli in microfluidics.

Neuronal activities of the human brain responsible for cognitive features have been theorized through several animal models that exhibited various complementary spatial learning modes by generating a flexible repertoire of behavioral strategies. However, for such studies associated with a neurodegenerative disease, which can be further manipulated to provide therapeutic strategies, the animal models employed in their developmental stages have been preferred over the adult ones. This pilot work was incepted to underscore the spatial memory capabilities that strengthened the intricate mechanism of memory acquisition potential in one of the low-order evolutionarily conserved species, such as zebrafish larvae. Initially, a reliable and more easily reproducible microfluidic platform integrating simple and intricate paths was designed to learn and test the spatial information in zebrafish larvae of 4-6 d.p.f. under non-invasive acoustic stimuli. Further, to acquire spatial information as the representation of spatial memory formation in zebrafish larvae, the acoustic startle responses were evaluated by quantifying various dynamic behaviors under distinct operating parameters. After significant conditioning sessions, the spatial memory was tested by employing variable 'freezing'. By the end of the 30 min-long test session, 6 d.p.f. larvae were found to exhibit the highest value of freezing of approximately 43% and 20% in the short and long paths, respectively. Even though a substantial rate of memory loss was observed, it can be envisaged to serve several behavioral strategies that process the dynamic cognitive memory among distinct spatiotemporal environments. Further, the proposed behavioral paradigm had the advantage of being more adaptable and reliably replicable by other researchers. As a consequence, different hypotheses can be readily tested to generate more reproducible findings towards distinct neurobehavioral characteristics. Therefore, the proposed paradigm for the consolidation of spatial memory based on the non-invasive spatial avoidance strategies could provide an enduring framework of reference for behavioral studies using zebrafish larvae.

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.

Emerging integrated SERS-microfluidic devices for analysis of cancer-derived small extracellular vesicles.

Cancer-derived small extracellular vesicles (sEVs) are specific subgroups of lipid bilayer vesicles secreted from cancer cells to the extracellular environment. They carry distinct biomolecules (e.g., proteins, lipids and nucleic acids) from their parent cancer cells. Therefore, the analysis of cancer-derived sEVs can provide valuable information for cancer diagnosis. However, the use of cancer-derived sEVs in clinics is still limited due to their small size, low amounts in circulating fluids, and heterogeneous molecular features, making their isolation and analysis challenging. Recently, microfluidic technology has gained great attention for its ability to isolate sEVs in minimal volume. In addition, microfluidics allows the isolation and detection of sEVs to be integrated into a single device, offering new opportunities for clinical application. Among various detection techniques, surface-enhanced Raman scattering (SERS) has emerged as a promising candidate for integrating with microfluidic devices due to its ultra-sensitivity, stability, rapid readout, and multiplexing capability. In this tutorial review, we start with the design of microfluidics devices for isolation of sEVs and introduce the key factors to be considered for the design, and then discuss the integration of SERS and microfluidic devices by providing descriptive examples of the currently developed platforms. Lastly, we discuss the current limitations and provide our insights for utilising integrated SERS-microfluidics to isolate and analyse cancer-derived sEVs in clinical settings.