Publications

Conventional gut-on-chip (GOC) models typically represent the epithelial layer of the gut tissue, neglecting other important components such as the stromal compartment and the extracellular matrix (ECM) that play crucial roles in maintaining intestinal barrier integrity and function. These models often employ hard, flat porous membranes for cell culture, thus failing to recapitulate the soft environment and complex 3D architecture of the intestinal mucosa. Alternatively, hydrogels have been recently introduced in GOCs as ECM analogs to support the co-culture of intestinal cells in in vivo-like configurations, and thus opening new opportunities in the organ-on-chip field. In this work, we present an innovative GOC device that includes a 3D bioprinted hydrogel channel replicating the intestinal villi architecture containing both the epithelial and stromal compartments of the gut mucosa. The bioprinted hydrogels successfully support both the encapsulation of fibroblasts and their co-culture with intestinal epithelial cells under physiological flow conditions. Moreover, we successfully integrated electrodes into the microfluidic system to monitor the barrier formation in real time via transepithelial electrical resistance measurements.

JTD Keywords: A-chip, Bioprinted, Caco-2, Cells, Culture, Gut-on-a-chip, Hydrogels, Impedance spectroscopy, Integrated electrodes, Intestinal barrier, Intestinal mucos, Model

The performance of single-chain polymeric nanoparticles (SCPNs) in biomedical applications highly depends on their conformational stability in cellular environments. Until now, such stability studies are limited to 2D cell culture models, which do not recapitulate the 3D tumor microenvironment well. Here, a microfluidic tumor-on-a-chip model is introduced that recreates the tumor milieu and allows in-depth insights into the diffusion, cellular uptake, and stability of SCPNs. The chip contains Matrigel/collagen-hyaluronic acid as extracellular matrix (ECM) models and is seeded with cancer cell MCF7 spheroids. With this 3D platform, it is assessed how the polymer's microstructure affects the SCPN's behavior when crossing the ECM, and evaluates SCPN internalization in 3D cancer cells. A library of SCPNs varying in microstructure is prepared. All SCPNs show efficient ECM penetration but their cellular uptake/stability behavior depends on the microstructure. Glucose-based nanoparticles display the highest spheroid uptake, followed by charged nanoparticles. Charged nanoparticles possess an open conformation while nanoparticles stabilized by internal hydrogen bonding retain a folded structure inside the tumor spheroids. The 3D microfluidic tumor-on-a-chip platform is an efficient tool to elucidate the interplay between polymer microstructure and SCPN's stability, a key factor for the rational design of nanoparticles for targeted biological applications.© 2024 The Authors. Small Methods published by Wiley-VCH GmbH.

JTD Keywords: 3d cancer cell uptake, Cancer cells, Cell culture, Cell uptake, Cellular uptake, Diseases, Ecm penetration, Extracellular matrices, Extracellular matrix penetration, Functional polymers, Hydrogen bonds, Medical applications, Microfluidics, Microstructure, Nanoparticles, Polymeric nanoparticles, Scpns, Single chains, Single-chain polymeric nanoparticle, Stability, Tumor-on-a-chip, Tumors

Neuroblastoma (NB) is a highly vascularized pediatric tumor arising from undifferentiated neural crest cells early in life, exhibiting both traditional endothelial-cell-driven vasculature and an intriguing alternative vasculature. The alternative vasculature can arise from cancer cells undergoing transdifferentiation into tumor-derived endothelial cells (TEC), a trait associated with drug resistance and tumor relapse. The lack of effective treatments targeting NB vasculature primarily arises from the challenge of establishing predictive in vitro models that faithfully replicate the alternative vasculature phenomenon. In this study, we aim to recreate the intricate vascular system of NB in an in vitro context, encompassing both types of vascularization, by developing a novel neuroblastoma-on-a-chip model. We designed a collagen I/fibrin-based hydrogel closely mirroring NB's physiological composition and tumor stiffness. This biomaterial created a supportive environment for the viability of NB and endothelial cells. Implementing a physiological shear stress value, aligned with the observed range in arteries and capillaries, within the microfluidic chip facilitated the successful development of vessel-like structures and triggered transdifferentiation of NB cells into TECs. The vascularized neuroblastoma-on-a-chip model introduced here presents a promising and complementary strategy to animal-based research with a significant capacity for delving into NB tumor biology and vascular targeting therapy.

JTD Keywords: 3d tumor model, Angiogenesis, Endothelial-cells, Microfluidic device, Neuroblastoma, Origi, Transdifferentiation, Tumor, Tumor-derived endothelial cells, Tumor-on-a-chip, Vasculature

Functional precision medicine (FPM) has emerged as a new approach to improve cancer treatment. Despite its potential, FPM assays present important limitations such as the number of cells and trained personnel required. To overcome these impediments, here we describe a novel microfluidic platform that can be used to perform FPM assays, optimizing the use of primary cancer cells and simplifying the process by using microfluidics to automatize the process.© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.

JTD Keywords: Bioassay, Biological assay, Cancer treatment, Functional assays, Lab-on-a-chip devices, Microfluidics, Personalized medicine, Precision medicine

Degradable polymeric micelles are promising drug delivery systems due to their hydrophobic core and responsive design. When applying micellar nanocarriers for tumor delivery, one of the bottlenecks encountered in vivo is the tumor tissue barrier: crossing the dense mesh of cells and the extracellular matrix (ECM). Sometimes overlooked, the extracellular matrix can trap nanoformulations based on charge, size, and hydrophobicity. Here, we used a simple design of a microfluidic chip with two types of ECM and MCF7 spheroids to allow high-throughput screening of the interactions between biological interfaces and polymeric micelles. To demonstrate the applicability of the chip, a small library of fluorescently labeled polymeric micelles varying in their hydrophilic shell and hydrophobic core forming blocks was studied. Three widely used hydrophilic shells were tested and compared, namely, poly(ethylene glycol), poly(2-ethyl-2-oxazoline), and poly(acrylic acid), along with two enzymatically degradable dendritic hydrophobic cores (based on hexyl or nonyl end groups). Using ratiometric imaging of unimer:micelle fluorescence and FRAP inside the chip model, we obtained the local assembly state and dynamics inside the chip. Notably, we observed different micelle behaviors in the basal lamina ECM, from avoidance of the ECM structure to binding of the poly(acrylic acid) formulations. Binding to the basal lamina correlated with higher uptake into MCF7 spheroids. Overall, we proposed a simple microfluidic chip containing dual ECM and spheroids for the assessment of the interactions of polymeric nanocarriers with biological interfaces and evaluating nanoformulations' capacity to cross the tumor tissue barrier.

JTD Keywords: Extracellular matrix, Microfluidics, Nanoparticle mobility, Polymeric micelles, Tumor-on-a-chip

Many neurodegenerative diseases are identified but their causes and cure are far from being well-known. The problem resides in the complexity of the neural tissue and its location which hinders its easy evaluation. Although necessary in the drug discovery process, in vivo animal models need to be reduced and show relevant differences with the human tissues that guide scientists to inquire about other possible options which lead to in vitro models being explored. From organoids to organ-on-a-chips, 3D models are considered the cutting-edge technology in cell culture. Cell choice is a big parameter to take into consideration when planning an in vitro model and cells capable of mimicking both healthy and diseased tissue, such as induced pluripotent stem cells (iPSC), are recognized as good candidates. Hence, we present a critical review of the latest models used to study neurodegenerative disease, how these models have evolved introducing microfluidics platforms, 3D cell cultures, and the use of induced pluripotent cells to better mimic the neural tissue environment in pathological conditions.

JTD Keywords: 3d in vitro models, bioprinting, ipsc cell culture, microfluidic device, 3d in vitro models, Bioprinting, Blood-brain-barrier, Cerebral organoids, Culture model, Endothelial-cells, Expression profile, Extracellular-matrix, Ipsc cell culture, Microfluidic device, Neurodegenerative diseases, On-a-chip, Pluripotent stem-cells, Shear-stress, Substrate stiffness

Alzheimer's disease is characterized by a combination of several neuropathological hallmarks, such as extracellular aggregates of beta amyloid (Aβ). Numerous alternatives have been studied for inhibiting Aβ aggregation but, at this time, there are no effective treatments available. Here, we developed the tri-component nanohybrid system AuNPs@POM@PEG based on gold nanoparticles (AuNPs) covered with polyoxometalates (POMs) and polyethylene glycol (PEG). In this work, AuNPs@POM@PEG demonstrated the inhibition of the formation of amyloid fibrils, showing a 75% decrease in Aβ aggregation in vitro. As it is a potential candidate for the treatment of Alzheimer's disease, we evaluated the cytotoxicity of AuNPs@POM@PEG and its ability to cross the blood-brain barrier (BBB). We achieved a stable nanosystem that is non-cytotoxic below 2.5 nM to human neurovascular cells. The brain permeability of AuNPs@POM@PEG was analyzed in an in vitro microphysiological model of the BBB (BBB-on-a-chip), containing 3D human neurovascular cell co-cultures and microfluidics. The results show that AuNPs@POM@PEG was able to cross the brain endothelial barrier in the chip and demonstrated that POM does not affect the barrier integrity, giving the green light to further studies into this system as a nanotherapeutic.

JTD Keywords: beta-amyloid, blood-brain barrier organ-on-a-chip, cellular uptake, citrate, cytotoxicity, electrocatalytic reduction, gold nanoparticles, hypothesis, nanorods, polyoxometalates, size, stability, surface, Alzheimers-disease, Blood–brain barrier organ-on-a-chip, Gold nanoparticles, Nanovehicle, Polyoxometalates, Β-amyloid

JTD Keywords: Cancer, Organ-on-a-chip / lab-on-a-chip / organoids and ex vivo models

JTD Keywords: Enabling technologies, Organ-on-a-chip / lab-on-a-chip / organoids and ex vivo models

JTD Keywords: Organ-on-a-chip / lab-on-a-chip / organoids and ex vivo models, Vascular systems / vascularisation and heart

Muscular dystrophies are a heterogeneous group of highly debilitating diseases that result in muscle atrophy and weakness. The lack of suitable cellular and animal models that reproduce specific aspects of their pathophysiology is one of the reasons why there are no curative treatments for these disorders. This highlights a considerable gap between current laboratory models and clinical practice. We strongly believe that organs-on-chip could help to fill this gap. Organs-on-chip, and in particular muscles-on-chip, are microfluidic devices that integrate functional skeletal muscle tissues. Biosensors in these systems allow monitoring of muscle homeostasis or drug responses in situ. This Perspective outlines the potential of organs-on-chip as advanced models for muscular dystrophies, as well as the current challenges and future opportunities for this technology.© 2023. Published by The Company of Biologists Ltd.

JTD Keywords: cell, tissue, Animals, Lab-on-a-chip devices, Muscle, skeletal, Muscular dystrophies, Skeletal-muscle

JTD Keywords: beta-cell function, organ-on-a-chip, pancreatic islets, plasmonic biosensors, responses, skeletal muscle, tissue engineering, Microfluidics, Type-2 diabetic-patients

The lack of predictive models that mimic the blood-brain barrier (BBB) hinders the development of effective drugs for neurodegenerative diseases. Animal models behave differently from humans, are expensive and have ethical constraints. Organ-on-a-chip (OoC) platforms offer several advantages to resembling physiological and pathological conditions in a versatile, reproducible, and animal-free manner. In addition, OoC give us the possibility to incorporate sensors to determine cell culture features such as trans-endothelial electrical resistance (TEER). Here, we developed a BBB-on-a-chip (BBB-oC) platform with a TEER measurement system in close distance to the barrier used for the first time for the evaluation of the permeability performance of targeted gold nanorods for theranostics of Alzheimer's disease. GNR-PEG-Ang2/D1 is a therapeutic nanosystem previously developed by us consisting of gold nanorods (GNR) functionalized with polyethylene glycol (PEG), angiopep-2 peptide (Ang2) to overcome the BBB and the D1 peptide as beta amyloid fibrillation inhibitor, finally obtaining GNR-PEG-Ang2/D1 which showed to be useful for disaggregation of the amyloid in in vitro and in vivo models. In this work, we evaluated its cytotoxicity, permeability, and some indications of its impact on the brain endothelium by employing an animal-free device based on neurovascular human cells.In this work, we fabricated a BBB-oC with human astrocytes, pericytes and endothelial cells and a TEER measuring system (TEER-BBB-oC) integrated at a micrometric distance of the endothelial barrier. The characterization displayed a neurovascular network and the expression of tight junctions in the endothelium. We produced GNR-PEG-Ang2/D1 and determined its non-cytotoxic range (0.05-0.4 nM) for plated cells included in the BBB-oC and confirmed its harmless effect at the highest concentration (0.4 nM) in the microfluidic device. The permeability assays revealed that GNR-PEG-Ang2/D1 cross the BBB and this entry is facilitated by Ang2 peptide. Parallel to the permeability analysis of GNR-PEG-Ang2/D1, an interesting behavior of the TJs expression was observed after its administration probably related to the ligands on the nanoparticle surface.BBB-oC with a novel TEER integrated setup which allow a correct read-out and cell imaging monitoring was proven as a functional and throughput platform to evaluate the brain permeability performance of nanotherapeutics in a physiological environment with human cells, putting forward a viable alternative to animal experimentation.© 2023. The Author(s).

JTD Keywords: alzheimer disease (ad), cell-culture, cytotoxicity, endothelial-cells, gold nanoparticles, microfluidic platform, model, organ-on-a-chip (ooc), peptide, tight junction, trans-endothelial electrical resistance (teer), transport, Alzheimer disease (ad), Blood-brain barrier (bbb), Blood-brain-barrier, Blood–brain barrier (bbb), Gold nanoparticles, Organ-on-a-chip (ooc), Trans-endothelial electrical resistance (teer)

In recent years, metaplasmonic biosensors have emerged as a novel counterpart of well-established plasmonic biosensors based on thin metallic layers. Metaplasmonic biosensors offer high potential for sensor miniaturiza-tion, extreme sensitivity biosensing, and high multiplexing capabilities with detection methods free of coupling optical elements. These capabilities make metaplasmonic biosensors highly attractive for Point-of-Care and handled/portable devices or novel On-Chip devices; as a result, it has increased the number of prototypes and potential applications that emerged during the last years. One of the main challenges to achieving fully operative devices is the achievement of high-throughput biointerfaces for sensitive and selective biodetection in complex media. Despite the superior surface sensitivity achieved by metaplasmonic sensors compared to conventional plasmonic sensors based on metallic thin films, the main limitations to achieving high-throughput and multiplexed biosensing usually are associated with the sensitivity and selectivity of the bioin-terface and, as a consequence, their application to the direct analysis of real complex samples. This graphical review discusses the potential challenges and capabilities of different biofunctionalization strategies, biorecog-nition elements, and antifouling strategies to achieve scalable and high-throughput metaplasmonic biosensing for Point-of-Care devices and bioengineering applications like Organs-On-Chip.

JTD Keywords: Biointerfaces, Biosensing, Biosensors, Cell culture monitoring, Metaplasmonic, Nanoplasmonic, Organ-on-chip, Point-of-care

The in vitro biological characterization of biomaterials is largely based on static cell cultures. However, for highly reactive biomaterials such as calcium-deficient hydroxyapatite (CDHA), this static environment has limitations. Drastic alterations in the ionic composition of the cell culture medium can negatively affect cell behavior, which can lead to misleading results or data that is difficult to interpret. This challenge could be addressed by a microfluidics-based approach (i.e. on-chip), which offers the opportunity to provide a continuous flow of cell culture medium and a potentially more physiologically relevant microenvironment. The aim of this work was to explore microfluidic technology for its potential to characterize CDHA, particularly in the context of inflammation. Two different CDHA substrates (chemically identical, but varying in microstructure) were integrated on-chip and subsequently evaluated. We demonstrated that the on-chip environment can avoid drastic ionic alterations and increase protein sorption, which was reflected in cell studies with RAW 264.7 macrophages. The cells grown on-chip showed a high cell viability and enhanced proliferation compared to cells maintained under static conditions. Whereas no clear differences in the secretion of tumor necrosis factor alpha (TNF-α) were found, variations in cell morphology suggested a more anti-inflammatory environment on-chip. In the second part of this study, the CDHA substrates were loaded with the drug Trolox. We showed that it is possible to characterize drug release on-chip and moreover demonstrated that Trolox affects the TNF-α secretion and morphology of RAW 264.7 ​cells. Overall, these results highlight the potential of microfluidics to evaluate (bioactive) biomaterials, both in pristine form and when drug-loaded. This is of particular interest for the latter case, as it allows the biological characterization and assessment of drug release to take place under the same dynamic in vitro environment.© 2022 The Authors.

JTD Keywords: alpha-tocopherol, antioxidant, biomaterials, calcium phosphate cement, culture, delivery, drug release, in vitro, in-vitro, ion, macrophage, on-chip, release, tool, Biomaterial, Calcium phosphate cement, Calcium-phosphate cements, Drug release, In vitro, Macrophage, On-chip

Precision medicine is starting to incorporate functional assays to evaluate anticancer agents on patient-isolated tissues or cells to select for the most effective. Among these new technologies, dynamic BH3 profiling (DBP) has emerged and extensively been used to predict treatment efficacy in different types of cancer. DBP uses synthetic BH3 peptides to measure early apoptotic events ('priming') and anticipate therapy-induced cell death leading to tumor elimination. This predictive functional assay presents multiple advantages but a critical limitation: the cell number requirement, that limits drug screening on patient samples, especially in solid tumors. To solve this problem, we developed an innovative microfluidic-based DBP (µDBP) device that overcomes tissue limitations on primary samples. We used microfluidic chips to generate a gradient of BIM BH3 peptide, compared it with the standard flow cytometry based DBP, and tested different anticancer treatments. We first examined this new technology's predictive capacity using gastrointestinal stromal tumor (GIST) cell lines, by comparing imatinib sensitive and resistant cells, and we could detect differences in apoptotic priming and anticipate cytotoxicity. We then validated µDBP on a refractory GIST patient sample and identified that the combination of dactolisib and venetoclax increased apoptotic priming. In summary, this new technology could represent an important advance for precision medicine by providing a fast, easy-to-use and scalable microfluidic device to perform DBP in situ as a routine assay to identify the best treatment for cancer patients.© 2022. The Author(s).

JTD Keywords: biomarkers, cancer drugs, chemotherapy, chip, models, platform, sensitivity, strategy, tumor-cells, Precision medicine

Organ-on-chip platforms combined with high-throughput sensing technology allow bridging gaps in information presented by 2D cultures modeled on static microphysiological systems. While these platforms do not aim to replicate whole organ systems with all physiological nuances, they try to mimic relevant structural, physiological, and functional features of organoids and tissues to best model disease and/or healthy states. The advent of this platform has not only challenged animal testing but has also presented the opportunity to acquire real-time, high-throughput data about the pathophysiology of disease progression by employing biosensors. Biosensors allow monitoring of the release of relevant biomarkers and metabolites as a result of physicochemical stress. It, therefore, helps conduct quick lead validation to achieve personalized medicine objectives. The organ-on-chip industry is currently embarking on an exponential growth trajectory. Multiple pharmaceutical and biotechnology companies are adopting this technology to enable quick patient-specific data acquisition at substantially low costs.

JTD Keywords: A-chip, Biosensor, Biosensors, Cancer, Cells, Culture, Disease models, Epithelial electrical-resistance, Hydrogel, Microfabrication, Microphysiological systems, Models, Niches, Organ-on-a-chips, Platform

Acute Respiratory Distress Syndrome is one of the more common fatal complications in COVID-19, characterized by a highly aberrant inflammatory response. Pre-clinical models to study the effect of cell therapy and anti-inflammatory treatments have not comprehensively reproduced the disease due to its high complexity. This work presents a novel physiomimetic in vitro model for Acute Respiratory Distress Syndrome using lung extracellular matrix-derived hydrogels and organ-on-a-chip devices. Monolayres of primary alveolar epithelial cells were cultured on top of decellullarized lung hydrogels containing primary lung mesenchymal stromal cells. Then, cyclic stretch was applied to mimic breathing, and an inflammatory response was induced by using a bacteriotoxin hit. Having simulated the inflamed breathing lung environment, we assessed the effect of an anti-inflammatory drug (i.e., dexamethasone) by studying the secretion of the most relevant inflammatory cytokines. To better identify key players in our model, the impact of the individual factors (cyclic stretch, decellularized lung hydrogel scaffold, and the presence of mesenchymal stromal cells) was studied separately. Results showed that developed model presented a more reduced inflammatory response than traditional models, which is in line with what is expected from the response commonly observed in patients. Further, from the individual analysis of the different stimuli, it was observed that the use of extracellular matrix hydrogels obtained from decellularized lungs had the most significant impact on the change of the inflammatory response. The developed model then opens the door for further in vitro studies with a better-adjusted response to the inflammatory hit and more robust results in the test of different drugs or cell therapy.

JTD Keywords: alveolar epithelial cells, ards, extracellular matrix, hydrogels, inflammation, lung-on-a-chip, Acute lung injury, Alveolar epithelial cells, Ards, Dexamethasone, Epithelial-mesenchymal transition, Extracellular matrix, Extracellular-matrix, Hydrogels, Inflammation, Lung-on-a-chip, Mesenchymal stromal cells, Oxygen, Stem-cells

This manuscript presents portable and low cost electronic system for specific point-of-use dielectrophoresis applications. The system is composed of two main modules: a) a multiphase generator based on a Class E amplifier, which provides 4 sinusoidal signals (0°, 90°, 180°, 270°) at 1 MHz with variable output voltage up to 10 Vpp (Vm) and an output driving current of 1 A; and b) a dielectrophoresis-based microfluidic chip containing two interdigitated electrodes. The system has been validated by concentrating Escherichia Coli at 1 MHz while applying a continuous flow of 5 ?L/min. Device functionalities were verified under different conditions achieving a 83% trapping efficiency in the best case. © Springer International Publishing Switzerland 2014.

JTD Keywords: bacteria, emulsifier, enterobacter, exopolysaccharide, Bacteria, Bacteria concentrations, Biochemical engineering, Cell concentrator, Class e amplifier, Class-e amplifier, Device functionality, Dielectrophoresis, Electronic equipment, Electronics, Electrophoresis, Emulsifier, Enterobacter, Escherichia coli, Exopolysaccharide, Inter-digitated electrodes, Lab-on-a-chip (loc), Low cost, Low costs, Low-cost electronics, Medical computing, Monosaccharide, Portable device, Power amplifiers, Trapping efficiencies

Nowadays, liquid biopsy represents one of the most promising techniques for early diagnosis, monitoring, and therapy screening of cancer. This novel methodology includes, among other techniques, the isolation, capture, and analysis of circulating tumor cells (CTCs). Nonetheless, the identification of CTC from whole blood is challenging due to their extremely low concentration (1-100 per ml of whole blood), and traditional methods result insufficient in terms of purity, recovery, throughput and/or viability of the processed sample. In this context, the development of microfluidic devices for detecting and isolating CTCs offers a wide range of new opportunities due to their excellent properties for cell manipulation and the advantages to integrate and bring different laboratory processes into the microscale improving the sensitivity, portability, reducing cost and time. This chapter explores current and recent microfluidic approaches that have been developed for the analysis and detection of CTCs, which involve cell capture methods based on affinity binding and label-free methods and detection based on electrical, chemical, and optical sensors. All the exposed technologies seek to overcome the limitations of commercial systems for the analysis and isolation of CTCs, as well as to provide extended analysis that will allow the development of novel and more efficient diagnostic tools.© 2022. The Author(s), under exclusive license to Springer Nature Switzerland AG.

JTD Keywords: cancer detection, cancer diagnosis, cancer-cells, capture, chip, circulating tumor cells, enrichment, liquid biopsy, microchannel, separation, ultra-fast, Cancer detection, Cancer diagnosis, Circulating tumor cells, Label-free isolation, Liquid biopsy, Microfluidics

Biosensors represent a powerful analytical tool for analyzing biomolecular interactions with the potential to achieve real-time quantitative analysis with high accuracy using low sample volumes, minimum sample pretreatment with high potential for the development of in situ and highly integrated monitoring platforms. Considering these advantages, their use in cell-culture systems has increased over the last few years. Between the different technologies for cell culture, organs-on-a-chip (OOCs) represent a novel technology that tries to mimic an organ's functionality by combining tissue engineering/organoid with microfluidics. Although there are still challenges to achieving OOC models with high organ mimicking relevance, these devices can offer effective models for drug treatment development by identifying drug targets, screening toxicity, and determining the potential effects of drugs in living beings. Consequently, in the future, we might replace animal studies by offering more ethical test models. Considering the relevance that different physiological and biochemical parameters have in the correct functionality of cells, sensing and biosensing platforms can offer an effective way for the real-time monitoring of physiological parameters and, in our opinion, more relevant, the secretion of biomarkers such as cytokines, growth factors, and others related with the influence of drugs or other types of stimulus in cell metabolism. Keeping this concept in mind, in this chapter, we focus on describing the potential use of sensors and biosensors in OOC devices to achieve fully integrated platforms that monitor physiological parameters and cell metabolism.© 2022. The Author(s), under exclusive license to Springer Nature Switzerland AG.

JTD Keywords: alignment, biosensors, cell, crystal microbalance biosensor, electrochemical biosensors, future, graphene oxide, label-free detection, organ-on-a-chip, oxygen, pre-clinical platforms, real-time analysis, screening, Biosensors, Organ-on-a-chip, Pre-clinical platforms, Screening, Sensors, Surface-plasmon resonance

Over the most recent decades, the development of new biological platforms to study disease progression and drug efficacy has been of great interest due to the high increase in the rate of neurodegenerative diseases (NDDs). Therefore, blood-brain barrier (BBB) as an organ-on-a-chip (OoC) platform to mimic brain-barrier performance could offer a deeper understanding of NDDs as well as a very valuable tool for drug permeability testing for new treatments. A very attractive improvement of BBB-oC technology is the integration of detection systems to provide continuous monitoring of biomarkers in real time and a fully automated analysis of drug permeably, rendering more efficient platforms for commercialization. In this Perspective, an overview of the main BBB-oC configurations is introduced and a critical vision of the BBB-oC platforms integrating electronic read out systems is detailed, indicating the strengths and weaknesses of current devices, proposing the great potential for biosensors integration in BBB-oC. In this direction, we name potential biomarkers to monitor the evolution of NDDs related to the BBB and/or drug cytotoxicity using biosensor technology in BBB-oC.

JTD Keywords: biosensors, blood−brain barrier (bbb), neurodegenerative diseases (ndds), organ-on-a-chip (ooc), Bbb, Biosensors, Blood-brain barrier (bbb), Electrical-resistance, Electrochemical biosensors, Endothelial-cells, In-vitro model, Matrix metalloproteinases, Mechanisms, Neurodegenerative diseases (ndds), Organ-on-a-chip (ooc), Permeability, Stress, Transendothelial electrical resistance (teer), Transepithelial, Transepithelial/transendothelial electrical resistance (teer), Transport

Diabetes mellitus is a significant public health problem worldwide. It encompasses a group of chronic disorders characterized by hyperglycemia, resulting from pancreatic islet dysfunction or as a consequence of insulin-producing ?-cell death. Organ-on-a-chip platforms have emerged as technological systems combining cell biology, engineering, and biomaterial technological advances with microfluidics to recapitulate a specific organ’s physiological or pathophysiological environment. These devices offer a novel model for the screening of pharmaceutical agents and to study a particular disease. In the field of diabetes, a variety of microfluidic devices have been introduced to recreate native islet microenvironments and to understand pancreatic ?-cell kinetics in vitro. This kind of platforms has been shown fundamental for the study of the islet function and to assess the quality of these islets for subsequent in vivo transplantation. However, islet physiological systems are still limited compared to other organs and tissues, evidencing the difficulty to study this “organ” and the need for further technological advances. In this review, we summarize the current state of islet-on-a-chip platforms that have been developed so far. We recapitulate the most relevant studies involving pancreatic islets and microfluidics, focusing on the molecular and cellular-scale activities that underlie pancreatic ?-cell function.

JTD Keywords: pancreatic islets, Diabetes, Microchips, Microfluidics

Microphysiological systems (MPS) or organs-on-chips (OoC) can emulate the physiological functions of organs in vitro and are effective tools for determining human drug responses in preclinical studies. However, the analysis of MPS has relied heavily on optical tools, resulting in difficulties in real-time and high spatial resolution imaging of the target cell functions. In this study, the role of scanning probe microscopy (SPM) as an analytical tool for MPS is evaluated. An access hole is made in a typical MPS system with stacked microchannels to insert SPM probes into the system. For the first study, a simple vascular model composed of only endothelial cells is prepared for SPM analysis. Changes in permeability and local chemical flux are quantitatively evaluated during the construction of the vascular system. The morphological changes in the endothelial cells after flow stimulation are imaged at the single-cell level for topographical analysis. Finally, the possibility of adapting the permeability and topographical analysis using SPM for the intestinal vascular system is further evaluated. It is believed that this study will pave the way for an in situ permeability assay and structural analysis of MPS using SPM.

JTD Keywords: cell, electrochemical microscopy, membrane-permeability, microphysiological systems, organs-chips, platform, scanning electrochemical microscopy, scanning ion conductance microscopy, scanning probe microscopy, shear-stress, surface-topography, Ion conductance microscopy, Microphysiological systems, Organs-chips, Scanning electrochemical microscopy, Scanning ion conductance microscopy, Scanning probe microscopy

The creation of cardiac tissue models for preclinical testing is still a non-solved problem in drug discovery, due to the limitations related to thein vitroreplication of cardiac tissue complexity. Among these limitations, the difficulty of mimicking the functional properties of the myocardium due to the immaturity of the used cells hampers the obtention of reliable results that could be translated into human patients.In vivomodels are the current gold standard to test new treatments, although it is widely acknowledged that the used animals are unable to fully recapitulate human physiology, which often leads to failures during clinical trials. In the present work, we present a microfluidic platform that aims to provide a range of signaling cues to immature cardiac cells to drive them towards an adult phenotype. The device combines topographical electrospun nanofibers with electrical stimulation in a microfabricated system. We validated our platform using a co-culture of neonatal mouse cardiomyocytes and cardiac fibroblasts, showing that it allows us to control the degree of anisotropy of the cardiac tissue inside the microdevice in a cost-effective way. Moreover, a 3D computational model of the electrical field was created and validated to demonstrate that our platform is able to closely match the distribution obtained with the gold standard (planar electrode technology) using inexpensive rod-shaped biocompatible stainless-steel electrodes. The functionality of the electrical stimulation was shown to induce a higher expression of the tight junction protein Cx-43, as well as the upregulation of several key genes involved in conductive and structural cardiac properties. These results validate our platform as a powerful tool for the tissue engineering community due to its low cost, high imaging compatibility, versatility, and high-throughput configuration capabilities.

JTD Keywords: bioreactor, cardiac tissue engineering, cardiomyocytes, electrospinning, fabrication, fibroblasts, heart-on-a-chip, heart-tissue, in vitro models, myocardium, orientation, platform, scaffolds, Cardiac tissue engineering, Electrospinning, Field stimulation, Heart-on-a-chip, In vitro models, Microphysiological system

Combining microfluidics technology with machine learning represents an innovative approach to conduct massive quantitative cell behavior study and implement smart decision-making systems in support of clinical diagnostics. The spleen plays a key-role in rare hereditary hemolytic anemia (RHHA), being the organ responsible for the premature removal of defective red blood cells (RBCs). The goal is to adapt the physiological spleen filtering strategy for in vitro study and monitoring of blood diseases through RBCs shape analysis. Then, a microfluidic device mimicking the slits of the spleen red pulp area and video data analysis are combined for the characterization of RBCs in RHHA. This microfluidic unit is designed to evaluate RBC deformability by maintaining them fixed in planar orientation, allowing the visual inspection of RBC's capacity to restore their original shape after crossing microconstrictions. Then, two cooperative learning approaches are used for the analysis: the majority voting scheme, in which the most voted label for all the cell images is the class assigned to the entire video; and the maximum sum of scores to decide the maximally scored class to assign. The proposed platform shows the capability to discriminate healthy controls and patients with an average efficiency of 91%, but also to distinguish between RHHA subtypes, with an efficiency of 82%.

JTD Keywords: chip, disease, Red-blood-cell

Tumor vessel-on-a-chip systems have attracted the interest of the cancer research community due to their ability to accurately recapitulate the multiple dynamic events of the metastatic cascade. Vessel-on-a-chip microfluidic platforms have been less utilized for investigating the distinctive features and functional heterogeneities of tumor-derived vascular networks. In particular, vascular tumors are characterized by the massive formation of thrombi and severe bleeding, a rare and life-threatening situation for which there are yet no clear therapeutic guidelines. This is mainly due to the lack of technological platforms capable of reproducing these characteristic traits of the pathology in a simple and well-controlled manner. Herein, we report the fabrication of a versatile tumor vessel-on-a-chip platform to reproduce, investigate, and characterize the massive formation of thrombi and hemorrhage on-chip in a fast and easy manner. Despite its simplicity, this method offers multiple advantages to recapitulate the pathophysiological events of vascular tumors, and therefore, may find useful applications in the field of vascular-related diseases, while at the same time being an alternative to more complex approaches. © 2021 by the authors. Licensee MDPI, Basel, Switzerland.

JTD Keywords: in vitro model, microfluidics, organ-on-chip, vascular tumor, vessel, In vitro model, Microfluidics, Organ-on-chip, Vascular tumor, Vessel

Organ-on-a-chip (OOC) devices offer new approaches for metabolic disease modeling and drug discovery by providing biologically relevant models of tissues and organs in vitro with a high degree of control over experimental variables for high-content screening applications. Yet, to fully exploit the potential of these platforms, there is a need to interface them with integrated non-labeled sensing modules, capable of monitoring, in situ, their biochemical response to external stimuli, such as stress or drugs. In order to meet this need, we aim here to develop an integrated technology based on coupling a localized surface plasmon resonance (LSPR) sensing module to an OOC device to monitor the insulin in situ secretion in pancreatic islets, a key physiological event that is usually perturbed in metabolic diseases such as type 2 diabetes (T2D). As a proof of concept, we developed a biomimetic islet-on-a-chip (IOC) device composed of mouse pancreatic islets hosted in a cellulose-based scaffold as a novel approach. The IOC was interfaced with a state-of-the-art on-chip LSPR sensing platform to monitor the in situ insulin secretion. The developed platform offers a powerful tool to enable the in situ response study of microtissues to external stimuli for applications such as a drug-screening platform for human models, bypassing animal testing.

JTD Keywords: biosensor, cytoarchitecture, dna hybridization, gelatin, in situ insulin monitoring, langerhans, lspr sensors, microfluidic device, organ-on-a-chip, parallel, platform, scaffold, Animals, Biosensing techniques, Diabetes mellitus, type 2, Drug discovery, Drug evaluation, preclinical, Human pancreatic-islets, Humans, In situ insulin monitoring, Insulin secretion, Insulins, Lab-on-a-chip devices, Lspr sensors, Oligonucleotide array sequence analysis, Organ-on-a-chip, Surface plasmon resonance

Tissue barriers play a crucial role in human physiology by establishing tissue compartmentalization and regulating organ homeostasis. At the interface between the extracellular matrix (ECM) and flowing fluids, epithelial and endothelial barriers are responsible for solute and gas exchange. In the past decade, microfluidic technologies and organ-on-chip devices became popular as in vitro models able to recapitulate these biological barriers. However, in conventional microfluidic devices, cell barriers are primarily grown on hard polymeric membranes within polydimethylsiloxane (PDMS) channels that do not mimic the cell-ECM interactions nor allow the incorporation of other cellular compartments such as stromal tissue or vascular structures. To develop models that accurately account for the different cellular and acellular compartments of tissue barriers, researchers have integrated hydrogels into microfluidic setups for tissue barrier-on-chips, either as cell substrates inside the chip, or as self-contained devices. These biomaterials provide the soft mechanical properties of tissue barriers and allow the embedding of stromal cells. Combining hydrogels with microfluidics technology provides unique opportunities to better recreate in vitro the tissue barrier models including the cellular components and the functionality of the in vivo tissues. Such platforms have the potential of greatly improving the predictive capacities of the in vitro systems in applications such as drug development, or disease modeling. Nevertheless, their development is not without challenges in their microfabrication. In this review, we will discuss the recent advances driving the fabrication of hydrogel microfluidic platforms and their applications in multiple tissue barrier models.

JTD Keywords: hydrogel, microfabrication, microfluidics, organ-on-chip, tissue barrier, Hydrogel, Microfabrication, Microfluidics, Organ-on-chip, Tissue barrier

Non-alcoholic fatty liver affects about 25% of global adult population. On the long-term, it is associated with extra-hepatic compliances, multiorgan failure, and death. Various invasive and non-invasive methods are employed for its diagnosis such as liver biopsies, CT scan, MRI, and numerous scoring systems. However, the lack of accuracy and reproducibility represents one of the biggest limitations of evaluating the effectiveness of drug candidates in clinical trials. Organ-on-chips (OOC) are emerging as a cost-effective tool to reproduce in vitro the main NAFLD’s pathogenic features for drug screening purposes. Those platforms have reached a high degree of complexity that generate an unprecedented amount of both structured and unstructured data that outpaced our capacity to analyze the results. The addition of artificial intelligence (AI) layer for data analysis and interpretation enables those platforms to reach their full potential. Furthermore, the use of them do not require any ethic and legal regulation. In this review, we discuss the synergy between OOC and AI as one of the most promising ways to unveil potential therapeutic targets as well as the complex mechanism(s) underlying NAFLD.

JTD Keywords: artificial intelligence, extra-hepatic outcome, nafld, organ-on-a-chip, Artificial intelligence, Extra-hepatic outcome, Nafld, Organ-on-a-chip

© 2021 by the authors. Licensee MDPI, Basel, Switzerland. Microfabrication and Polydimethylsiloxane (PDMS) soft-lithography techniques became popular for microfluidic prototyping at the lab, but even after protocol optimization, fabrication is yet a long, laborious process and partly user-dependent. Furthermore, the time and money required for the master fabrication process, necessary at any design upgrade, is still elevated. Digital Manufacturing (DM) and Rapid-Prototyping (RP) for microfluidics applications arise as a solution to this and other limitations of photo and soft-lithography fabrication techniques. Particularly for this paper, we will focus on the use of subtractive DM techniques for Organ-on-a-Chip (OoC) applications. Main available thermoplastics for microfluidics are suggested as material choices for device fabrication. The aim of this review is to explore DM and RP technologies for fabrication of an OoC with an embedded membrane after the evaluation of the main limitations of PDMS soft-lithography strategy. Different material options are also reviewed, as well as various bonding strategies. Finally, a new functional OoC device is showed, defining protocols for its fabrication in Cyclic Olefin Polymer (COP) using two different RP technologies. Different cells are seeded in both sides of the membrane as a proof of concept to test the optical and fluidic properties of the device.

JTD Keywords: digital manufacturing, microfluidics, organ on a chip, rapid prototyping, Digital manufacturing, Microfluidics, Organ on a chip, Rapid prototyping

© 2020 American Chemical Society. The performance of supramolecular nanocarriers as drug delivery systems depends on their stability in the complex and dynamic biological media. After administration, nanocarriers are challenged by physiological barriers such as shear stress and proteins present in blood, endothelial wall, extracellular matrix, and eventually cancer cell membrane. While early disassembly will result in a premature drug release, extreme stability of the nanocarriers can lead to poor drug release and low efficiency. Therefore, comprehensive understanding of the stability and assembly state of supramolecular carriers in each stage of delivery is the key factor for the rational design of these systems. One of the main challenges is that current 2D in vitro models do not provide exhaustive information, as they fail to recapitulate the 3D tumor microenvironment. This deficiency in the 2D model complexity is the main reason for the differences observed in vivo when testing the performance of supramolecular nanocarriers. Herein, we present a real-time monitoring study of self-assembled micelles stability and extravasation, combining spectral confocal microscopy and a microfluidic cancer-on-a-chip. The combination of advanced imaging and a reliable 3D model allows tracking of micelle disassembly by following the spectral properties of the amphiphiles in space and time during the crucial steps of drug delivery. The spectrally active micelles were introduced under flow and their position and conformation continuously followed by spectral imaging during the crossing of barriers, revealing the interplay between carrier structure, micellar stability, and extravasation. Integrating the ability of the micelles to change their fluorescent properties when disassembled, spectral confocal imaging and 3D microfluidic tumor blood vessel-on-a-chip resulted in the establishment of a robust testing platform suitable for real-time imaging and evaluation of supramolecular drug delivery carrier's stability.

JTD Keywords: cancer-on-a-chip, complex, delivery, endothelial-cells, in-vitro, microfluidic, model, nanoparticle, penetration, shear-stress, stability, supramolecular, Cancer-on-a-chip, Cell-culture, Micelle, Microfluidic, Nanoparticle, Stability, Supramolecular

Human aging is accompanied by a relevant increase in age-associated chronic pathologies, including neurodegenerative and metabolic diseases. The appearance and evolution of numerous neurodegenerative diseases is paralleled by the appearance of intracellular and extracellular accumulation of misfolded proteins in affected brains. In addition, recent evidence suggests that most of these amyloid proteins can behave and propagate among neural cells similarly to infective prions. In order to improve understanding of the seeding and spreading processes of these “prion-like” amyloids, microfluidics and 3D lab-on-chip approaches have been developed as highly valuable tools. These techniques allow us to monitor changes in cellular and molecular processes responsible for amyloid seeding and cell spreading and their parallel effects in neural physiology. Their compatibility with new optical and biochemical techniques and their relative availability have increased interest in them and in their use in numerous laboratories. In addition, recent advances in stem cell research in combination with microfluidic platforms have opened new humanized in vitro models for myriad neurodegenerative diseases affecting different cellular targets of the vascular, muscular, and nervous systems, and glial cells. These new platforms help reduce the use of animal experimentation. They are more reproducible and represent a potential alternative to classical approaches to understanding neurodegeneration. In this review, we summarize recent progress in neurobiological research in “prion-like” protein using microfluidic and 3D lab-on-chip approaches. These approaches are driven by various fields, including chemistry, biochemistry, and cell biology, and they serve to facilitate the development of more precise human brain models for basic mechanistic studies of cell-to-cell interactions and drug discovery.

JTD Keywords: Lab-On-Chip, Amyloid propagation, Microfluidics, Fibril, Seeding, Spreading, Prion-like, Prionoid

Epithelial tissues are composed of layers of tightly connected cells shaped into complex three-dimensional (3D) structures such as cysts, tubules, or invaginations. These complex 3D structures are important for organ-specific functions and often create biochemical gradients that guide cell positioning and compartmentalization within the organ. One of the main functions of epithelia is to act as physical barriers that protect the underlying tissues from external insults. In vitro, epithelial barriers are usually mimicked by oversimplified models based on cell lines grown as monolayers on flat surfaces. While useful to answer certain questions, these models cannot fully capture the in vivo organ physiology and often yield poor predictions. In order to progress further in basic and translational research, disease modeling, drug discovery, and regenerative medicine, it is essential to advance the development of new in vitro predictive models of epithelial tissues that are capable of representing the in vivo-like structures and organ functionality more accurately. Here, we review current strategies for obtaining biomimetic systems in the form of advanced in vitro models that allow for more reliable and safer preclinical tests. The current state of the art and potential applications of self-organized cell-based systems, organ-on-a-chip devices that incorporate sensors and monitoring capabilities, as well as microfabrication techniques including bioprinting and photolithography, are discussed. These techniques could be combined to help provide highly predictive drug tests for patient-specific conditions in the near future.

JTD Keywords: 3D cell culture models, Biofabrication, Disease modeling, Drug screening, Epithelial barriers, Microengineered tissues, Organ-on-a-chip, Organoids

Suspended planar-array (SPA) chips embody millions of individual miniaturized arrays to work in extremely small volumes. Here, the basis of a robust methodology for the fabrication of SPA silicon chips with on-demand physical and chemical anisotropies is demonstrated. Specifically, physical traits are defined during the fabrication process with special focus on the aspect ratio, branching, faceting, and size gradient of the final chips. Additionally, the chemical attributes augment the functionality of the chips with the inclusion of complete coverage or patterns of selected biomolecules on the surface of the chips with contact printing techniques, offering an extremely high versatility, not only with the choice of the pattern shape and distribution but also in the choice of biomolecular inks to pattern. This approach increases the miniaturization of printed arrays in 3D structures by two orders of magnitude compared to those previously demonstrated. Finally, functional micrometric and sub-micrometric patterned features are demonstrated with an antibody binding assay with the recognition of the printed spots with labeled antibodies from solution. The selective addition of physical and chemical attributes on the suspended chips represents the basis for future biomedical assays performed within extremely small volumes.

JTD Keywords: Microcontact printing, Microparticles, Molecular multiplexing, Polymer pen lithography, Silicon chip technology

A novel suspended planar-array chips technology is described, which effectively allows molecular multiplexing using a single suspended chip to analyze extraordinarily small volumes. The suspended chips are fabricated by combining silicon-based technology and polymer-pen lithography, obtaining increased molecular pattern flexibility, and improving miniaturization and parallel production. The chip miniaturization is so dramatic that it permits the intracellular analysis of living cells.

JTD Keywords: Chip-in-a-cell, Suspended-arrays, Planar-arrays, Silicon chips, Single-cell analysis

Intermittent hypoxia (IH), a hallmark of obstructive sleep apnea (OSA), plays a critical role in the pathogenesis of OSA-associated morbidities, especially in the cardiovascular and respiratory systems. Oxidative stress and inflammation induced by IH are suggested as main contributors of end-organ dysfunction in OSA patients and animal models. Since the molecular mechanisms underlying these in vivo pathological responses remain poorly understood, implementation of experimental in vitro cell-based systems capable of inducing high-frequency IH would be highly desirable. Here, we describe the design, fabrication, and validation of a versatile chip for subjecting cultured cells to fast changes in gas partial pressure and to cyclic stretch. The chip is fabricated with polydimethylsiloxane (PDMS) and consists of a cylindrical well-covered by a thin membrane. Cells cultured on top of the membrane can be subjected to fast changes in oxygen concentration (equilibrium time ~6 s). Moreover, cells can be subjected to cyclic stretch at cardiac or respiratory frequencies independently or simultaneously. Rat bone marrow-derived mesenchymal stem cells (MSCs) exposed to IH mimicking OSA and cyclic stretch at cardiac frequencies revealed that hypoxia-inducible factor 1a (HIF-1a) expression was increased in response to both stimuli. Thus, the chip provides a versatile tool for the study of cellular responses to cyclical hypoxia and stretch.

JTD Keywords: Cell stretch, Hypoxia-inducible factor, Intermittent hypoxia, Lab-on-a-chip, Obstructive sleep apnea

Bacteria concentration and detection is time-consuming in regular microbiology procedures aimed to facilitate the detection and analysis of these cells at very low concentrations. Traditional methods are effective but often require several days to complete. This scenario results in low bioanalytical and diagnostic methodologies with associated increased costs and complexity. In recent years, the exploitation of the intrinsic electrical properties of cells has emerged as an appealing alternative approach for concentrating and detecting bacteria. The combination of dielectrophoresis (DEP) and impedance analysis (IA) in microfluidic on-chip platforms could be key to develop rapid, accurate, portable, simple-to-use and cost-effective microfluidic devices with a promising impact in medicine, public health, agricultural, food control and environmental areas. The present document reviews recent DEP and IA combined approaches and the latest relevant improvements focusing on bacteria concentration and detection, including selectivity, sensitivity, detection time, and conductivity variation enhancements. Furthermore, this review analyses future trends and challenges which need to be addressed in order to successfully commercialize these platforms resulting in an adequate social return of public-funded investments.

JTD Keywords: Bacteria, Dielectrophoresis, Impedance, Microfluidics, On-chip

Pharmaceutical drug screening and research into diseases call for significant improvement in the effectiveness of current in vitro models. Better models would reduce the likelihood of costly failures at later drug development stages, while limiting or possibly even avoiding the use of animal models. In this regard, promising advances have recently been made by the so-called "organ-on-chip" (OOC) technology. By combining cell culture with microfluidics, biomedical researchers have started to develop microengineered models of the functional units of human organs. With the capacity to mimic physiological microenvironments and vascular perfusion, OOC devices allow the reproduction of tissue- and organ-level functions. When considering drug testing, nephrotoxicity is a major cause of attrition during pre-clinical, clinical, and post-approval stages. Renal toxicity accounts for 19% of total dropouts during phase III drug evaluation-more than half the drugs abandoned because of safety concerns. Mimicking the functional unit of the kidney, namely the nephron, is therefore a crucial objective. Here we provide an extensive review of the studies focused on the development of a nephron-on-chip device.

JTD Keywords: Disease model, Drug discovery, Kidney, Nephron-on-chip, Organ-on-chip

We describe a novel continuous-flow cell concentrator micro-device based on dielectrophoresis (DEP), and its associated custom-made control unit. The performances of a classical interdigitated metal electrode-based DEP microfluidic device and this enhanced version, that includes insulator-based pole structures, were compared using the same setup. Escherichia coli (E. coli) samples were concentrated at several continuous flows and the device's trapping efficiencies were evaluated by exhaustive cell counts. Our results show that pole structures enhance the retention up to 12.6%, obtaining significant differences for flow rates up to 20 μl/min, when compared to an equivalent classical interdigitated electrodes setup. In addition, we performed a subsequent proteomic analysis to evaluate the viability of the biological samples after the long exposure to the actuating electrical field. No E. coli protein alteration in any of the two systems was observed.

JTD Keywords: Concentrator, Dielectrophoresis, Escherichia coli, Lab- on- a- chip

This manuscript presents a portable and low cost electronic system for specific point-of-use dielectrophoresis applications. The system is composed of two main modules: a) a multiphase generator based on a Class E amplifier, which provides 4 sinusoidal signals (0°, 90°, 180°, 270°) at 1 MHz with variable output voltage up to 10 Vpp (Vm) and an output driving current of 1 A; and b) a dielectrophoresis-based microfluidic chip containing two interdigitated electrodes. The system has been validated by concentrating Escherichia coli (E. coli) at 1 MHz while applying a continuous flow of 5 µL/min. The device functionalities were verified under different conditions, achieving an 83% trapping efficiency when counter-phased signals are used.

JTD Keywords: Cell Concentrator, Class E amplifier, Dielectrophoresis, Electronics, Lab-on-a-chip (LOC), Low cost, Portable device

Constant evolution and improvements on areas such as tissue engineering, microfluidics and nanotechnology have made it possible to partially close the gap between conventional in vitro cell cultures and animal model-based studies. A step forward in this field concerns organ-on-chip technologies, capable of reproducing the most relevant physiological features of an organ in a microfluidic platform. In this work we have exploited the capabilities of laminar co-flow inside our biomimetic platform, the splenon-on-a-chip, in order to enhance cell culture inside its channels to better mimic the spleen's environment. © 14CBMS.

JTD Keywords: Cell culture, Co-flow, Laminar flow, Organ-on-a-chip, Spleen

Splenomegaly, albeit variably, is a landmark of malaria infection. Due to technical and ethical constraints, however, the role of the spleen in malaria remains vastly unknown. The spleen is a complex three-dimensional branched vasculature exquisitely adapted to perform different functions containing closed/rapid and open/slow microcirculations, compartmentalized parenchyma (red pulp, white pulp and marginal zone), and sinusoidal structure forcing erythrocytes to squeeze through interstitial slits before reaching venous circulation. Taking into account these features, we have designed and developed a newfangled microfluidic device of a human splenon-on-a-chip (the minimal functional unit of the red pulp facilitating blood-filtering and destruction of malarial-infected red blood cells). Our starting point consisted in translating splenon physiology to the most similar microfluidic network, mimicking the hydrodynamic behavior of the organ, to evaluate and simulate its activities, mechanics and physiological responses and, therefore, enable us to study biological hypotheses. Different physiological features have been translated into engineering elements that can be combined to integrate a biomimetic microfluidic spleen model. The device is fabricated in polydimethylsiloxane (PDMS), a biocompatible polymer, irreversibly bonded to glass. Microfluidics analyses have confirmed that 90% of the blood circulates through a fast-flow compartment whereas the remaining 10% circulates through a slow compartment, equivalently to what has been observed in a real spleen. Moreover, erythrocytes and reticulocytes going through the slow-flow compartment squeeze at the end of it through 2μm physical constraints resembling interstitial slits to reach the closed/rapid circulation.

JTD Keywords: Malaria, Microfluidics, Organ-on-a-chip, Spleen

This manuscript presents portable and low cost electronic system for specific point-of-use dielectrophoresis applications. The system is composed of two main modules: a) a multiphase generator based on a Class E amplifier, which provides 4 sinusoidal signals (0º, 90º, 180º, 270º) at 1 MHz with variable output voltage up to 10 Vpp (Vm) and an output driving current of 1 A; and b) a dielectrophoresis-based microfluidic chip containing two interdigitated electrodes. The system has been validated by concentrating Escherichia Coli at 1 MHz while applying a continuous flow of 5 μL/min. Device functionalities were verified under different conditions achieving a 83% trapping efficiency in the best case.

JTD Keywords: Cell Concentrator, Class E amplifier, Dielectrophoresis, Electronics, Lab-on-a-chip (LOC), Low cost, Portable device

The achievement of a higher degree of integration of components – especially micropumps and power sources – is a challenge currently being pursued to obtain portable and totally autonomous microfluidic devices. This paper presents the integration of a micro direct methanol fuel cell (mDMFC) in a microfluidic platform as a smart solution to provide both electrical and pumping power to a Lab-on-a-Chip system. In this system the electric power produced by the fuel cell is available to enable most of the functionalites required by the microfluidic chip, while the generated CO2 from the electrochemical reaction produces a pressure capable of pumping a liquid volume through a microchannel. The control of the fuel cell operating conditions allows regulation of the flow rate of a liquid sample through a microfluidic network. The relation between sample flow rate and the current generated by the fuel cell is practically linear, achieving values in the range of 4–18 mL min 1 while having an available power between 1–4 mW. This permits adjusting the desired flow rate for a given application by controlling the fuel cell output conditions and foresees a fully autonomous analytical Lab-on-a-Chip in which the same device would provide the electrical power to a detection module and at the same time use the CO2 pumping action to flow the required analytes through a particular microfluidic design.

JTD Keywords: micro direct methanol fuel cell (mDMFC), Lab-on-a-chip (LOC), Microfluidic device

Biosensors' research filed has clearly been changing towards the production of multifunctional and innovative design concepts to address the needs related with sensitivity and selectivity of the devices. More recently, waveguide biosensors, that do not require any label procedure to detect biomolecules adsorbed on its surface, have been pointed out as one of the most promising technologies for the production of biosensing devices with enhanced performance. Moreover the combination of optical and electrochemical measurements through the integration of transparent and conducting oxides in the multilayer structures can greatly enhance the biosensors' sensitivity. Furthermore, the integration of polymeric substrates may bring powerful advantages in comparison with silicon based ones. The biosensors will have a lower production costs being possible to disposable them after use ("one use sensor chip"). This research work represents a preliminary study about the influence of substrate temperature on the overall properties of ITO thin films deposited by DC magnetron sputtering onto 0,5 mm thick PMMA sheets.

JTD Keywords: ITO thin films, PMMA sheets, Waveguide biosensing devices, Biosensing devices, Conducting oxides, Dc magnetron sputtering, Electrochemical measurements, Enhanced performance, Innovative design, ITO thin films, Multilayer structures, Overall properties, PMMA sheets, Polymeric substrate, Production cost, Sensor chips, Silicon-based, Substrate temperature, Biosensors, Deposition, Design, Film preparation, Optical multilayers, Thin films, Vapor deposition, Waveguides, Substrates

The continuous flow focussing and manipulation of particles and cells are important factors in microfluidic applications for performing accurate and reproducible procedures downstream. Many particle focussing methods require complex setups or channel designs that can limit the process and its applications. Here, we present diamagnetic repulsion as a simple means of focussing objects in continuous flow, based only on their intrinsic properties without the requirement of any label. Diamagnetic polystyrene particles were suspended in a paramagnetic medium and pumped through a capillary between a pair of permanent magnets, whereupon the particles were repelled by each magnet into the central axis of the capillary, thus achieving focussing. By investigating this effect, we found that the focussing was greatly enhanced with (i) increased magnetic susceptibility of the medium, (ii) reduced flow rate of the suspension, (iii) increased particle size, and (iv) increased residence time in the magnetic field. Furthermore, we applied diamagnetic repulsion to the flow focussing of living, label-free HaCaT cells.

JTD Keywords: Feeble magnetic substances, On-chip, Blood-cells, Microfluidic device, Separation, Field, Levitation, Magnetophoresis, Fractionation, Nanoparticles

Electrokinetic techniques are contact-free methods currently used in many applications, where precise handling of biological entities, such as cells, bacteria or nucleic acids, is needed. These techniques are based on the effect of electric fields on molecules suspended in a fluid, and the corresponding induced motion, which can be tuned according to some known physical laws and observed behaviours. Increasing interest on the application of such strategies in order to improve the detection of DNA strands has appeared during the recent decades. Classical electrode-based DNA electrochemical biosensors with combined electrokinetic techniques present the advantage of being able to improve the working electrode's bioactive part during their fabrication and also the hybridization yield during the sensor detection phase. This can be achieved by selectively manipulating, driving and directing the molecules towards the electrodes increasing the speed and yield of the floating DNA strands attached to them. On the other hand, this technique can be also used in order to make biosensors reusable, or reconfigurable, by simply inverting its working principle and pulling DNA strands away from the electrodes. Finally, the combination of these techniques with nanostructures, such as nanopores or nanochannels, has recently boosted the appearance of new types of electrochemical sensors that exploit the time-varying position of DNA strands in order to continuously scan these molecules and to detect their properties. This review gives an insight into the main forces involved in DNA electrokinetics and discusses the state of the art and uses of these techniques in recent years.

JTD Keywords: Electrochemical DNA biosensors, Lab-on-a-chip (LOC), Micro-total analysis systems (mu TAS), Nanopore

A hybrid microfluidic device that uses hydrodynamic forces to separate human plasma from blood cells has been designed and fabricated and the advantageous effects of temperature and flow rates are investigated in this paper. The blood separating device includes an inlet which is reduced by approximately 20 times to a small constrictor channel, which then opens out to a larger output channel with a small lateral channel for the collection of plasma. When tested the device separated plasma from whole blood using a wide range of flow rates, between 50 mu l min(-1) and 200 mu l min(-1), at the higher flow rates injected by hand and at temperatures ranging from 23 degrees C to 50 degrees C, the latter resulting in an increase in the cell-free layer of up to 250%. It was also tested continuously using between 5% and 40% erythrocytes in plasma and whole blood without blocking the channels or hemolysis of the cells. The mean percentage of plasma collected after separation was 3.47% from a sample of 1 ml. The percentage of cells removed from the plasma varied depending on the flow rate used, but at 37 degrees C ranged between 95.4 +/- 1% and 97.05 +/- 05% at 100 mu l min(-1) and 200 mu l min(-1), respectively. The change in temperature also had an effect on the number of cells removed from the plasma which was between 93.5 +/- 0.65% and 97.01 +/- 0.3% at 26.9 degrees C and 37 degrees C, respectively, using a flow rate of 100 mu l min(-1). Due to its ability to operate in a wide range of conditions, it is envisaged that this device can be used in in vitro 'lab on a chip' applications, as well as a hand-held point of care (POC) device.

JTD Keywords: On-a-chip, Cells, Viscosity, Membrane

In this paper, a multiple-channel extension of a dual-grating Coupler biosensor is presented as a Solution for the problem of resolving different selectivities, Usual when heterogeneous samples are analyzed. Several differently functionalized channels can perform quantitative analysis of competiting recognition events, Suppress shifts due to buffer changes and even monitorize drifts coming from the light Source. Here, the multiple-channel approach is developed and proven for a four-channel configuration, providing a resolution limit of 10(-5) Refractive index Units (RIU) and with an a potentially Unlimited scalability. Finally, a differential HSA recognition event is monitored, at both an IgG functionalized channel and at a blocked one.

JTD Keywords: Optical grating coupler, Multi-channel biorecognition, On-chip reference

We report a new strategy for immunochemical screening of small organic molecules based on the use of a hapten microarray. Using DNA-directed immobilization strategies, we have been able to convert a DNA chip into a hapten microarray by taking advantage of all the benefits of the structural and electrostatic homogeneous properties of DNA. The hapten microarray uses hapten-oligonucleotide probes instead of proteins, avoiding the limitations of preparing stochiometrically-defined protein-oligonucleotide bioconjugates. As proof of concept, we show here the development of a microarray for analysis of anabolic androgenic steroids. The microchip is able to detect several illegal substances with sufficient detectability to be used as a screening method, according to the regulations of the World Anti-Doping Agency for sport and the European Commision for food safety. The results that we show corroborate the universal possibilities of the DNA chip, and, in this case, they open the way to develop hapten microarrays for the immunochemical analysis of small organic molecules.

JTD Keywords: Anti-doping, DNA chip, DNA-directed immobilization (DDI), Fluorescence, Food safety, Hapten microarray, Immunochemical screening, Proof of concept, Small organic molecule, Steroid

Analytical devices able to perform accurate and fast automatic DNA detection or sequencing procedures have many potential benefits in the biomedical and environmental fields. The conversion of biological or biochemical responses into quantifiable optical, mechanical or electronic signals is achieved by means of biosensors. Most of these transducing elements can be miniaturized and incorporated into lab-on-a-chip devices, also known as Micro Total Analysis Systems. The use of multiple DNA biosensors integrated in these miniaturized laboratories, which perform several analytical operations at the microscale, has many cost and efficiency advantages. Tiny amounts of reagents and samples are needed and highly sensitive, fast and parallel assays can be done at low cost. A particular type of DNA biosensors are the ones used based on electrochemical principles. These sensors offer several advantages over the popular fluorescence-based detection schemes. The resulting signal is electrical and can be processed by conventional electronics in a very cheap and fast manner. Furthermore, the integration and miniaturization of electrochemical transducers in a microsystem makes easier its fabrication in front of the most common currently used detection method. In this review, different electrochemical DNA biosensors integrated in analytical microfluidic devices are discussed and some early stage commercial products based on this strategy are presented.

JTD Keywords: DNA, Electrochemical DNA biosensors, Electrochemistry, Lab-on-a-chip, Micro Total Analysis systems, Field-effect transistors, Sequence-specific detection, Chemical-analysis systems, Solid-state nanopores, Carbon nanotubes, Microfluidic device, Electrical detection, Hybridization, Molecules, Sensor

This article presents the fabrication and characterisation of a high-speed detection micro-Coulter counter with two-dimensional (2D) adjustable aperture and differential impedance detection. The developed device has been fabricated from biocompatible and transparent materials (polymer and glass) and uses the principle of hydrodynamic focusing in two dimensions. The use of a conductive solution for the sample flux and non-conductive solutions for the focalising fluxes provides an adjustable sample flow where particles are aligned and the resistive response concentrated, consequently enhancing the sensitivity and versatility of the device. High-speed counting of 20 mu m polystyrene particles and 5 mu m yeast cells with a rate of up to 1000 particles/s has been demonstrated. Two-dimensional focusing conditions have been used in devices with physical cross-sectional areas of 180 mu m x 65 mu m and 100 mu m x 43 mu m, respectively, in which particles resulted undetectable in the absence of focusing. The 2D-focusing conditions have provided, in addition, increased detection sensitivity by a factor of 1.6 as compared to 1 D-focusing conditions.

JTD Keywords: Impedance, Chip, Microfluidics