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Neural differentiation is crucial for advancing our understanding of the nervous system and developing treatments for neurological disorders. The advanced methods and the ability to manipulate the alignment, proliferation, and differentiation of stem cells are essential for studying neuronal development and synaptic interactions. However, the utilization of human induced pluripotent stem cells (iPSCs) for disease modeling of neurodegenerative conditions may be constrained by the prolonged duration and uncontrolled cell differentiation required for functional neural cell differentiation. Here, we developed a microfluidic chip to enhance the differentiation and maturation of specific neural lineages by placing aligned microelectrodes on the glass surface to regulate the neural differentiation of human iPSCs. The utilization of electrical stimulation (ES) in conjunction with neurotrophic factors (NF) significantly enhanced the efficiency in generating functional neurons from human iPSCs. We also observed that the simultaneous application of NF and ES to human iPSCs promoted their differentiation and maturation into functional neurons while increasing synaptic interactions. Our research demonstrated the effect of combining NF and ES on human iPSC-derived neural differentiation.

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A new perspective suggests that a dynamic bidirectional communication system, often referred to as the microbiome-gut-brain axis, exists among the gut, its microbiome, and the central nervous system (CNS). This system may influence brain health and various brain-related diseases, especially in the realms of neurodevelopmental and neurodegenerative conditions. However, the exact mechanism is not yet understood. Metabolites or extracellular vesicles derived from microbes in the gut have the capacity to traverse the intestinal epithelial barrier or blood-brain barrier, gaining access to the systemic circulation. This phenomenon can initiate the physiological responses that directly or indirectly impact the CNS and its function. However, reliable and controllable tools are required to demonstrate the causal effects of gut microbial-derived substances on neurogenesis and neurodegenerative diseases. The integration of microfluidics enhances scientific research by providing advanced in vitro engineering models. In this study, we investigated the impact of microbe-derived metabolites and exosomes on neurodevelopment and neurodegenerative disorders using human induced pluripotent stem cells (iPSCs)-derived neurons in a gut-brain axis chip. While strain-specific, our findings indicate that both microbial-derived metabolites and exosomes exert the significant effects on neural growth, maturation, and synaptic plasticity. Therefore, our results suggest that metabolites and exosomes derived from microbes hold promise as potential candidates and strategies for addressing neurodevelopmental and neurodegenerative disorders.

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Cancer is a leading cause of the death worldwide. However, the conventional cancer therapy still suffers from several limitations, such as systemic side effects, poor efficacy, and patient compliance due to limited accessibility to the tumor site. To address these issues, the localized drug delivery system has emerged as a promising approach. In this study, we developed an iontophoresis-based transdermal drug delivery system (TDDS) controlled by a smartphone application for cancer treatment. Iontophoresis, a low-intensity electric current-based TDDS, enhances drug permeation across the skin to provide potential for localized drug delivery and minimize systemic side effects. The fundamental mechanism of our system was modeled using finite element analysis and its performance was corroborated through the flow-through skin permeation tests using a plastic-based microfluidic chip. The results of in vitro cell experiments and skin deposition tests successfully demonstrated that our smartphone-controlled iontophoresis system significantly enhanced the drug permeation for cancer treatment. Therefore, this hand-held smartphone-based iontophoresis TDDS could be a powerful tool for self-administrated anticancer drug delivery applications.

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The proposed electro-responsive hydrogel has great benefit for transdermal drug delivery system (TDDS) applications. To improve the physical or chemical properties of hydrogels, a number of researchers have previously studied the mixing efficiencies of the blended hydrogels. However, few studies have focused on improving the electrical conductivity and drug delivery of the hydrogels. We developed a conductive blended hydrogel by mixing alginate with gelatin methacrylate (GelMA) and silver nanowire (AgNW). We demonstrated that and the tensile strength of blended hydrogels were increased by a factor of 1.8 by blending GelMA and the electrical conductivity was enhanced by a factor of 18 by the addition of AgNW. Furthermore, the GelMA-alginate-AgNW (Gel-Alg-AgNW) blended hydrogel patch enabled on-off controllable drug release, indicating 57% doxorubicin release in response to electrical stimulation (ES) application. Therefore, this electro-responsive blended hydrogel patch could be useful for smart drug delivery applications.

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Since the outbreak of coronavirus 2019 (COVID-19), detection technologies have been attracting a great deal of attention in molecular diagnosis applications. In particular, the droplet digital PCR (ddPCR) has become a promising tool as it offers absolute quantification of target nucleic acids with high specificity and sensitivity. In recent years, the combination of the isothermal amplification strategies has made ddPCR a popular method for on-site testing by enabling amplification at a constant temperature. However, the current isothermal ddPCR assays are still challenging due to inherent non-specific amplification. In this paper, we present a multiplexed droplet digital recombinase polymerase amplification (MddRPA) with precise initiation of the reaction. First, the reaction temperature and dynamic range of reverse transcription (RT) and RPA were characterized by real-time monitoring of fluorescence intensities. Using a droplet-based microfluidic chip, the master mix and the initiator were fractionated and rapidly mixed within well-confined droplets. Due to the high heat transfer and mass transfer of the droplets, the precise initiation of the amplification was enabled and the entire assay could be conducted within 30 min. The concentrations of target RNA in the range from 5 copies per µL to 2500 copies per µL could be detected with high linearity (R2 > 0.999). Furthermore, the multiplexed detection of three types of human coronaviruses was successfully demonstrated with high specificity (>96%). Finally, we compared the performance of the assay with a commercial RT-qPCR system using COVID-19 clinical samples. The MddRPA assay showed a 100% concordance with the RT-qPCR results, indicating its reliability and accuracy in detecting SARS-CoV-2 nucleic acids in clinical samples. Therefore, our MddRPA assay with rapid detection, precise quantification, and multiplexing capability would be an interesting method for molecular diagnosis of viral infections.

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Since coronavirus disease 2019 (COVID-19) pandemic rapidly spread worldwide, there is an urgent demand for accurate and suitable nucleic acid detection technology. Although the conventional threshold-based algorithms have been used for processing images of droplet digital polymerase chain reaction (ddPCR), there are still challenges from noise and irregular size of droplets. Here, we present a combined method of the mask region convolutional neural network (Mask R-CNN)-based image detection algorithm and Gaussian mixture model (GMM)-based thresholding algorithm. This novel approach significantly reduces false detection rate and achieves highly accurate prediction model in a ddPCR image processing. We demonstrated that how deep learning improved the overall performance in a ddPCR image processing. Therefore, our study could be a promising method in nucleic acid detection technology.

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Metabolism, is a complex process involving the gut and the liver tissue, is difficult to be reproduced in vitro with conventional single cell culture systems. To tackle this challenge, we developed a gut-liver-axis chip consisting of the gut epithelial cell chamber and three-dimensional (3D) uniform-sized liver spheroid chamber. Two cell culture chamber compartments were separated with a porous membrane to prevent microorganisms from passing through the chamber. When the hepG2 spheroids cultured with microbiota-derived metabolites, we observed the changes in the physiological function of hepG2 spheroids, showing that the albumin and urea secretion activity of liver spheroids was significantly enhanced. Additionally, the functional validation of hepG2 spheroids treated with microbiota-derived exosome was evaluated that the treatment of the microbiota-derived exosome significantly enhanced albumin and urea in hepG2 spheroids in a gut-liver axis chip. Therefore, this gut-liver axis chip could be a potentially powerful co-culture platform to study the interaction of microbiota and host cells.

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The multi-stimuli responsive drug delivery system has recently attracted attention in cancer treatments, since it can reduce several side effects and enhance cancer therapeutic efficacy. Herein, we present the intracellular antioxidant (glutathione, GSH), enzyme (hyaluronidase, HAase), and hydrogen peroxide (H2O2) triggered mesoporous organo-silica (MOS) nanocomposites for multi-modal treatments via chemo-, photothermal, and photodynamic cancer therapies. A MOS nanoparticle was synthesized by two-types of precursors, tetraethyl orthosilicate (TEOS) and bis[3-(triethoxysilyl)propyl] tetrasulfide (BTES), providing large-sized mesopores and disulfide bonds cleavable by GSH. Additionally, we introduced a new ß-cyclodextrin-hyaluronic acid (CDHA) gatekeeper system, enabling nanocomposites to form the specific interaction with the ferrocene (Fc) molecule, control the drug release by the HAase and H2O2 environment, as well as provide the targeting ability against the CD44-overexpressing melanoma (B16F10) cells. Indocyanine green (ICG) and doxorubicin (Dox) were loaded in the MOS-Fc-CDHA (ID@MOS-Fc-CDHA) nanocomposites, allowing for hyperthermia and cytotoxic reactive oxygen species (ROS) under an 808 nm NIR laser irradiation. Therefore, we demonstrated that the ID@MOS-Fc-CDHA nanocomposites were internalized to the B16F10 cells via the CD44 receptor-mediated endocytosis, showing the controlled drug release by GSH, HAase, and H2O2 to enhance the cancer therapeutic efficacy via the synergistic chemo-, photothermal, and photodynamic therapy effect.

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A three-dimensional (3D) tumor spheroid model plays a critical role in mimicking tumor microenvironments in vivo. However, the conventional culture methods lack the ability to manipulate the 3D tumor spheroids in a homogeneous manner. To address this limitation, we developed a microfluidic-based droplet system for drug screening applications. We used a tree-shaped gradient generator to control the cell density and encapsulate the cells within uniform-sized droplets to generate a 3D gradient-sized tumor spheroid. Using this microfluidic-based droplet system, we demonstrated the high-throughput generation of uniform 3D tumor spheroids containing various cellular ratios for the analysis of the anti-cancer drug cytotoxicity. Consequently, this microfluidic-based gradient droplet generator could be a potentially powerful tool for anti-cancer drug screening applications.

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For rapid detection of the COVID-19 infection, the digital polymerase chain reaction (dPCR) with higher sensitivity and specificity has been presented as a promising method of point-of-care testing (POCT). Unlike the conventional real-time PCR (qPCR), the dPCR system allows absolute quantification of the target DNA without a calibration curve. Although a number of dPCR systems have previously been reported, most of these previous assays lack multiplexing capabilities. As different variants of COVID-19 have rapidly emerged, there is an urgent need for highly specific multiplexed detection systems. Additionally, the advances in the Internet of Things (IoT) technology have enabled the onsite detection of infectious diseases. Here, we present an IoT-integrated multiplexed dPCR (IM-dPCR) system involving sample compartmentalization, DNA amplification, fluorescence imaging, and quantitative analysis. This IM-dPCR system comprises three modules: a plasmonic heating-based thermal cycler, a multi-color fluorescence imaging set-up, and a firmware control module. Combined with a custom-developed smartphone application built on an IoT platform, the IM-dPCR system enabled automatic processing, data collection, and cloud storage. Using a self-priming microfluidic chip, 9 RNA groups (e.g., H1N1, H3N2, IFZ B, DENV2, DENV3, DENV4, OC43, 229E, and NL63) associated with three infectious diseases (e.g., influenza, dengue, and human coronaviruses) were analyzed with higher linearity (>98%) and sensitivity (1 copy per µL). The IM-dPCR system exhibited comparable analytical accuracy to commercial qPCR platforms. Therefore, this IM-dPCR system plays a crucial role in the onsite detection of infectious diseases.

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Polymerase chain reaction has gained attention since the outbreak of novel coronavirus in 2019. Due to its high specificity and capability for early detection, it is considered a standard method for the diagnosis of infectious diseases. However, the conventional thermocyclers used for nucleic acid amplification are not suitable for point-of-care testing applications, as they require expensive instruments, high-power consumption, and a long turnaround time. To suppress the widespread of the pandemic, there is an urgent need for the development of a rapid, inexpensive, and portable thermal cycler. Therefore, in this paper, we present a conductive silver/carbon fiber film-based thermal cycler with low power consumption (<5 W), efficient heating (~4.5 °C/s), low cost (

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The precise manipulation of the neural stem cell (NSC)-derived neural differentiation is still challenging, and there is a technological barrier to regulate the axonal regeneration in a controlled manner. Here, we developed a microfluidic chip integrated with a microelectrode array as an axonal guidance platform. The microfluidic electrode array chip consisted of two compartments and a bridge microchannel that could isolate and guide the axons. We demonstrated that the NSCs were largely differentiated into neural cells as the electric field was applied to the microfluidic electrode array chip. We also confirmed the synergistic effects of the electrical stimulation (ES) and neurotrophic factor (NF) on axonal outgrowth. This microfluidic electrode array chip can serve as a central nervous system (CNS) model for axonal injury and regeneration. Therefore, it could be a potentially powerful tool for an in vitro model of the axonal regeneration.

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The intestinal microbiome affects a number of biological functions of the organism. Although the animal model is a powerful tool to study the relationship between the host and microbe, a physiologically relevant in vitro human intestinal system has still unmet needs. Thus, the establishment of an in vitro living cell-based system of the intestine that can mimic the mechanical, structural, absorptive, transport and pathophysiological properties of the human intestinal environment along with its commensal bacterial strains can promote pharmaceutical development and potentially replace animal testing. In this paper, we present a microfluidic-based gut model which allows co-culture of human and microbial cells to mimic the gastrointestinal structure. The gut microenvironment is recreated by flowing fluid at a low rate (21 µL/h) over the microchannels. Under these conditions, we demonstrated the capability of gut-on-a-chip to recapitulate in vivo relevance epithelial cell differentiation including highly polarized epithelium, mucus secretion, and tight membrane integrity. Additionally, we observed that the co-culture of damaged epithelial layer with the probiotics resulted in a substantial responded recovery of barrier function without bacterial overgrowth in a gut-on-a-chip. Therefore, this gut-on-a-chip could promote explorations interaction with host between microbe and provide the insights into questions of fundamental research linking the intestinal microbiome to human health and disease.

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Near-infrared (NIR) light-mediated photothermal therapy (PTT) and photodynamic therapy (PDT) have widely been used for cancer treatment applications. However, a number of limitations (e.g., low NIR absorption capacity of photothermal agents, insufficient loading efficiency of photosensitive molecules) have hindered the widespread use of NIR-mediated cancer therapy. Therefore, we developed a mesoporous silica-coated reduced graphene oxide (rGO) nanocomposite that could provide a high encapsulation rate of indocyanine green (ICG) and enhance PTT/PDT efficiency in vitro and in vivo. The ICG-encapsulated nanocomposite not only enhances the photothermal effect but also generates a large number of tumor toxic reactive oxygen species (ROS). By conjugation of polyethylene glycol (PEG) with folic acid (FA) as a tumor targeting moiety, we confirmed that ICG-encapsulated mesoporous silica (MS)-coated rGO nanocomposite (ICG@MS-rGO-FA) exhibited high colloidal stability and intracellular uptake in folate receptor-expressing CT-26 colorectal cancer cells. Upon NIR laser irradiation, this ICG@MS-rGO-FA nanocomposite induced the apoptosis of only CT-26 cells via enhanced PTT and PDT effects without any damage to normal cells. Furthermore, the ICG@MS-rGO-FA nanocomposite revealed satisfactory tumor targeting and biocompatibility in CT-26 tumor-bearing mice, thereby enhancing the therapeutic effects of PTT and PDT in vivo. Therefore, this tumor-targeted ICG@MS-rGO-FA nanocomposite shows a great potential for phototherapy applications.

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Blended hydrogels play an important role in enhancing the properties (e.g., mechanical properties and conductivity) of hydrogels. In this study, we generated a conductive blended hydrogel, which was achieved by mixing gelatin methacrylate (GelMA) with collagen, and silver nanowire (AgNW). The ratio of GelMA, collagen and AgNW was optimized and was subsequently gelated by ultraviolet light (UV) and heat. The scanning electron microscope (SEM) image of the conductive blended hydrogels showed that collagen and AgNW were present in the GelMA hydrogel. Additionally, rheological analysis indicated that the mechanical properties of the conductive GelMA-collagen-AgNW blended hydrogels improved. Biocompatibility analysis confirmed that the human umbilical vein endothelial cells (HUVECs) encapsulated within the three-dimensional (3D), conductive blended hydrogels were highly viable. Furthermore, we confirmed that the molecule in the conductive blended hydrogel was released by electrical stimuli-mediated structural deformation. Therefore, this conductive GelMA-collagen-AgNW blended hydrogel could be potentially used as a smart actuator for drug delivery applications.

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Nanoparticle-based nucleic acid conjugates (NP-NACs) hold great promise for theragnostic (diagnostic and therapeutic) applications. However, several limitations have hindered the realization of their full potential in the clinical treatment of cancer and other diseases. In diagnosis, NP-NACs, combined with conventional optical sensing systems, have been applied for cancer detection in vitro, but low signal-to-noise ratios limit their broad in vivo applications. Meanwhile, the efficiency of NP-NAC-mediated cancer therapies has been limited through the adaptation of alternative pro-survival pathways in cancer cells. The recent emergence of personalized and precision medicine has outlined the importance of both accurate diagnosis and efficient therapeutics in a single platform. As such, we report the controlled assembly of hybrid graphene oxide/gold nanoparticle-based cancer-specific NACs (Au@GO NP-NACs) for multimodal imaging and combined therapeutics. Our developed Au@GO NP-NACs shows excellent surface-enhanced Raman scattering (SERS)-mediated live-cell cancer detection and multimodal synergistic cancer therapy through the use of photothermal, genetic, and chemotherapeutic strategies. Synergistic and selective killing of cancer cells were then demonstrated by using in vitro microfluidic models and nine different cancer cell lines by further incorporating near-infrared photothermal hyperthermia, a Topoisomerase II anti-cancer drug, and cancer targeting peptides. Moreover, with distinctive advantages of the Au@GO NP-NACs for cancer theragnostics, we further demonstrated precision cancer treatment through the detection of cancer cells in vivo using SERS followed by efficient ablation of the tumor. Therefore, our Au@GO NP-NACs could pave a new road for the advanced theragnostics of cancer as well as many other diseases.

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We developed copper sulfide (CuS)/reduced graphene oxide (rGO)-poly (ethylene glycol) (PEG) nanocomposites for photothermal bonding of a polymethyl methacrylate (PMMA)-based plastic lab-on-a-chip. The noncontact photothermal bonding of PMMA-based plastic labs-on-chip plays an important role in improving the stability and adhesion at a high-temperature as well as minimizing the solution leakage from microchannels when connecting two microfluidic devices. The CuS/rGO-PEG nanocomposites were used to bond a PMMA-based plastic lab-on-a-chip in a short time with a high photothermal effect by a near-infrared (NIR) laser irradiation. After the thermal bonding process, a gap was not generated in the PMMA-based plastic lab-on-a-chip due to the low viscosity and density of the CuS/rGO-PEG nanocomposites. We also evaluated the physical and mechanical properties after the thermal bonding process, showing that there was no solution leakage in PMMA-based plastic lab-on-a-chip during polymerase chain reaction (PCR) thermal cycles. Therefore, the CuS/rGO-PEG nanocomposite could be a potentially useful nanomaterial for non-contact photothermal bonding between the interfaces of plastic module lab-on-a-chip.

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In this study, potential protection of chlorophyll a from illumination and oxidation-induced decomposition has been examined using C-phycocyanin (C-PC) aqueous medium. Photo-oxidation resistance of chlorophyll a was monitored in various aqueous media using ultraviolet-visible spectroscopy and direct-infusion atmospheric pressure chemical ionization mass spectrometry analysis. The spectroscopy results showed that chlorophyll a in C-PC medium experienced the lowest rate of conversion to its derivatives; thus, it was demonstrated that chlorophyll a was mostly intact in the C-PC medium. Furthermore, the C-PC treated with chlorophyll a showed the lowest concentrations of malondialdehyde, and chlorophyll a in C-PC medium did not cause serious damage to human liver cells in vitro after intensive illumination. Therefore, we propose a new method of protecting chlorophyll a from photodegradation and oxidation using C-PC aqueous medium.

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The combination therapy based on multifunctional nanocomposites has been considered as a promising approach to improve cancer therapeutic efficacy. Herein, we report targeted multi-functional poly(N-isopropylacrylamide) (PNIPAM)-based nanocomposites for synergistic chemo-photothermal therapy toward breast cancer cells. To increase the transition temperature, acrylic acid (AAc) was added in synthetic process of PNIPAM, showing that the intrinsic lower critical solution temperature was changed to 42 °C . To generate the photothermal effect under near-infrared (NIR) laser irradiation (808 nm), polypyrrole (ppy) nanoparticles were uniformly decorated in PNIPAM-AAc. Folic acid (FA), as a cancer targeting ligand, was successfully conjugated on the surplus carboxyl groups in PNIPAM network. The drug release of PNIPAM-ppy-FA nanocomposites was efficiently triggered in response to the temperature change by NIR laser irradiation. We also confirmed that PNIPAM-ppy-FA was internalized to MDA-MB-231 breast cancer cells by folate-receptor-mediated endocytosis and significantly enhanced cancer therapeutic efficacy with combination treatment of chemo-photothermal effects. Therefore, our work encourages further exploration of multi-functional nanocarrier agents for synergistic therapeutic approaches to different types of cancer cells.

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A miniaturized polymerase chain reaction (PCR) system is not only important for medical applications in remote areas of developing countries, but also important for testing at ports of entry during global epidemics, such as the current outbreak of the coronavirus. Although there is a large number of PCR sensor systems available for this purpose, there is still a lack of portable digital PCR (dPCR) heating systems. Here, we first demonstrated a portable plasmonic heating-based dPCR system. The device has total dimensions of 9.7 × 5.6 × 4.1 cm and a total power consumption of 4.5 W, allowing for up to 25 dPCR experiments to be conducted on a single charge of a 20 000 mAh external battery. The dPCR system has a maximum heating rate of 10.7 °C s-1 and maximum cooling rate of 8 °C s-1. Target DNA concentrations in the range from 101 ± 1.4 copies per µL to 260 000 ± 20 000 copies per µL could be detected using a poly(dimethylsiloxane) (PDMS) microwell membrane with 22 080 well arrays (20 µm diameter). Furthermore, the heating system was demonstrated using a mass producible poly(methyl methacrylate) PMMA microwell array with 8100 microwell arrays (80 µm diameter). The PMMA microwell array could detect a concentration from 12 ± 0.7 copies per µL to 25 889 ± 737 copies per µL.

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