Detection and monitoring of lactate and glucose levels in biological fluids and cell cultures are essential for understanding metabolic disorders. While electrochemical biosensors are commonly used, traditional enzymatic sensors face challenges related to stability, reproducibility, and cost. To address these limitations, we developed non-enzymatic sensors for lactate and glucose detection using nanostructured nickel oxide (NiO)–modified screen-printed carbon electrodes. The sensors were fabricated by drop-casting a NiO/Nafion/ethanol dispersion onto the working electrode, and their performance was evaluated using cyclic voltammetry and amperometry. Optimal sensitivity and linearity were achieved at a working potential of ∼0.5 V. The sensors exhibited linear responses for both lactate and glucose in the 0.1–5 mM range, with detection limits of 0.03 mM (lactate) and 0.025 mM (glucose), and sensitivities of 1.564 μA/mM (lactate) and 1.842 μA/mM (glucose) in 0.1 M NaOH–KCl electrolyte. To address glucose interference in lactate sensing, dual-sensing strategies were employed by varying Nafion concentration, applying differential potentials, or modifying the sensors with Prussian Blue to achieve selective detection. Validation against commercial lactate and glucose assay kits in cell culture medium showed good agreement, confirming the sensors’ accuracy. Finally, the sensor was integrated with a microfluidic chip, demonstrating its potential as a flow-through, enzyme-free metabolic sensor for future organ-on-a-chip applications.

A lab-in-a-fiber component was fabricated using an optical fiber and a fiber capillary. It was used in a test suspension of fluorescently labeled and unlabeled cells and enabled detection of the labeled cells. Subsequently the labeled cells were selectively collected via suction into the capillary. A novel sampling technique reduced photobleaching of the labeled cells, extending the measurement time. The collected cells remained viable for downstream analysis. This platform’s low fabrication cost, simplicity, compatibility with standard laboratory equipment, and capacity for fully automated cell capture highlights its potential for future applications in minimally invasive sample collection and point-of-care diagnostics. We demonstrate this LiF device to showcase the capability of optical fiber technology in creating low-cost, low-complexity cancer diagnostic devices. Furthermore, the LiF device holds promise for in vivo diagnostics, facilitating cell isolation and analysis.

The advancement of biopharmaceutical manufacturing, particularly continuous processing, has heightened the need for next-generation analytical tools approaching real-time monitoring of critical quality attributes (CQAs) and process parameters (CPPs). Current methods, primarily offline and labor-intensive, fail at delivering analytical information that can be used for process analytical technology (PAT) to control and optimize the manufacturing process, while also lacking the ability of multi-attribute monitoring, thus requiring a large number of samples (or sampling amount) to be collected. This work introduces the concept of PAT-on-a-chip, consisting of an integrated microfluidic platform designed to perform at-line analysis and characterization of cell culture samples in the context of monoclonal antibody (mAb) production. Specifically, a sample preparation-free miniaturized lectin-based assay was developed to measure levels of high mannose glycans and integrated with affinity-based assays to measure mAb titers and key impurities, namely Chinese hamster ovary (CHO) host cell proteins (HCP), within the same chip, resorting to a common colorimetric readout. The microfluidic chips were operated in a customized and integrated instrument comprising miniaturized photodiodes, connected to a graphical user interface for data recording and signal quantification. The PAT-on-a-chip unit allowed to achieve fit-for-purpose analyte quantification, while offering performance comparable to state-of-the-art offline analytical methods (Pearson R > 0.93), namely capillary electrophoresis with laser-induced fluorescence (CE-LIF) for glycan analysis, well plate immunoassays for CHO HCP and protein A HPLC for mAb titers, thus validating its potential to expand the modern PAT toolbox.

Nanoscale biological particles, such as lipoproteins (10–80 nm) or extracellular vesicles (30–200 nm), play pivotal roles in health and disease, including conditions like cardiovascular disorders and cancer. Their effective analysis is crucial for applications in diagnostics, quality control, and nanomedicine development. While elasto-inertial focusing offers a powerful method to manipulate particles without external fields, achieving consistent focusing of nanoparticles (<500 nm) has remained a challenge. In this study, elasto-inertial focusing of nanoparticles as small as 25 nm is experimentally demonstrated using straight high-aspect-ratio microchannels in a sheathless flow. Systematic investigations reveal the influence of channel width, particle size, viscoelastic concentration, and flow rate on focusing behavior. Additionally, through numerical simulations and experimental validation, insights are provided into particle migration dynamics and viscoelastic forces governing nanoparticle focusing. Finally, biological particles, including liposomes (90–140 nm), extracellular vesicles (100 nm), and lipoproteins (10–25 nm) is successfully focused, under optimized conditions, showcasing potential applications in medical diagnostics and targeted drug delivery. These findings mark a significant advancement toward size-based high-resolution particle separation, with implications for biomedicine and environmental sciences.

Laboratory automation effectively increases the throughput in sample analysis, reduces human errors in sample processing, as well as simplifies and accelerates the overall logistics. Automating diagnostic testing workflows in peripheral laboratories and also in near-patient settings -like hospitals, clinics and epidemic control checkpoints- is advantageous for the simultaneous processing of multiple samples to provide rapid results to patients, minimize the possibility of contamination or error during sample handling or transport, and increase efficiency. However, most automation platforms are expensive and are not easily adaptable to new protocols. Here, we address the need for a versatile, easy-to-use, rapid and reliable diagnostic testing workflow by combining open-source modular automation (Opentrons) and automation-compatible molecular biology protocols, easily adaptable to a workflow for infectious diseases diagnosis by detection on paper-based diagnostics. We demonstrated the feasibility of automation of the method with a low-cost Neisseria meningitidis diagnostic test that utilizes magnetic beads for pathogen DNA isolation, isothermal amplification, and detection on a paper-based microarray. In summary, we integrated open-source modular automation with adaptable molecular biology protocols, which was also faster and cheaper to perform in an automated than in a manual way. This enables a versatile diagnostic workflow for infectious diseases and we demonstrated this through a low-cost N. meningitidis test on paper-based microarrays.

The health hazards of micro- and nanoplastic contaminants in drinking water has recently emerged as an area of concern to policy makers and industry. Plastic contaminants range in size from micro- (5 mm to 1 μm) to nanoplastics (<1 μm). Microfluidics provides many tools for particle manipulation at the microscale, particularly in diagnostics and biomedicine, but has in general a limited capacity to process large volumes. Drinking water and environmental samples with low-level contamination of microplastics require processing of deciliter to liter sample volumes to achieve statistically relevant particle counts. Here, we introduce the EchoGrid, an acoustofluidics device for high throughput continuous flow particle enrichment into a robust array of particle clusters. The EchoGrid takes advantage of highly efficient particle capture through the integration of a micropatterned transducer for surface displacement-based acoustic trapping in a glass and polymer microchannel. Silica seed particles were used as anchor particles to improve capture performance at low particle concentrations and high flow rates. The device was able to maintain the silica grids at a flow rate of 50 mL/min. In terms of enrichment, the device is able to double the final pellet’s microplastic concentration every 78 s for 23 μm particles and every 51 s for 10 μm particles at a flow rate of 5 mL/min. In conclusion, we demonstrate the usefulness of the EchoGrid by capturing microplastics in challenging conditions, such as large sample volumes with low microparticle concentrations, without sacrificing the potential of integration with downstream analysis for environmental monitoring.

Micro- and nanoplastics have become increasingly relevant as contaminants to be monitored due to their potential health effects and environmental impact. Nanoplastics, in particular, have been shown to be difficult to detect in drinking water, requiring new capture technologies. In this work, we applied the acoustofluidic seed particle method to capture nanoplastics in an optimized, tilted grid of silica clusters even at the high flow rate of 5 mL/min. Moreover, we achieved, using this technique, the enrichment of nanoparticles ranging from 500 nm to 25 nm as a first in the field. We employed fluorescence to observe the enrichment profiles according to size, using a washing buffer flow at 0.5 mL/min, highlighting the size-dependent nature of the silica seed particle release of various sizes of nanoparticles. These results highlight the versatility of acoustic trapping for a wide range of nanoplastic particles and allow further study into the complex dynamics of the seed particle method at these size ranges. Moreover, with reproducible size-dependent washing curves, we provide a new window into the rate of nanoplastic escape in high-capacity acoustic traps, relevant to both environmental and biomedical applications.

The combination of flow elasticity and inertia has emerged as a viable tool for focusing and manipulating particles using microfluidics. Although there is considerable interest in the field of elasto-inertial microfluidics owing to its potential applications, research on particle focusing has been mostly limited to low Reynolds numbers (Re<1), and particle migration toward equilibrium positions has not been extensively examined. In this work, we thoroughly studied particle focusing on the dynamic range of flow rates and particle migration using straight microchannels with a single inlet high aspect ratio. We initially explored several parameters that had an impact on particle focusing, such as the particle size, channel dimensions, concentration of viscoelastic fluid, and flow rate. Our experimental work covered a wide range of dimensionless numbers (0.05 < Reynolds number < 85, 1.5 < Weissenberg number < 3800, 5 < Elasticity number < 470) using 3, 5, 7, and 10 µm particles. Our results showed that the particle size played a dominant role, and by tuning the parameters, particle focusing could be achieved at Reynolds numbers ranging from 0.2 (1 µL/min) to 85 (250 µL/min). Furthermore, we numerically and experimentally studied particle migration and reported differential particle migration for high-resolution separations of 5 µm, 7 µm and 10 µm particles in a sheathless flow at a throughput of 150 µL/min. Our work elucidates the complex particle transport in elasto-inertial flows and has great potential for the development of high-throughput and high-resolution particle separation for biomedical and environmental applications. (Figure presented.)

Aims: While enhanced tumor cell migration is a key process in the tumor dissemination, mechanistic insights into causal relationships between tumor cells and mechanical confinement are still limited. Here we combine the use of microfluidic platforms to characterize confined cell migration with genomic tools to systematically unravel the global signaling landscape associated with the migratory phenotype of breast cancer (BC) cells. Meterials and methods: The spontaneous migration capacity of seven BC cell lines was evaluated in 3D microfluidic devices and their migration capacity was correlated with publicly available molecular signatures. The role of identified signaling pathways on regulating BC migration capacity was determined by receptor stimulation through ligand binding or inhibition through siRNA silencing. Downstream effects on cell migration were evaluated in microfluidic devices, while the molecular changes were monitored by RT-qPCR. Key findings: Expression of 715 genes was correlated with BC cells migratory phenotype, revealing TNF-α as one of the top upstream regulators. Signal transduction experiments revealed that TNF-α stimulates the confined migration of triple negative, mesenchymal-like BC cells that are also characterized by high TNFR1 expression, but inhibits the migration of epithelial-like cells with low TNFR1 expression. TNFR1 was strongly associated with the migration capacity and triple-negative, mesenchymal phenotype. Downstream of TNF/TNFR1 signaling, transcriptional regulation of NFKB seems to be important in driving cell migration in confined spaces. Significance: TNF-α/TNFR1 signaling axis reveals as a key player in driving BC cells confined migration, emerging as a promising therapeutic strategy in targeting dissemination and metastasis of triple negative, mesenchymal BC cells.

Within the vast field of medical biotechnology, the biopharmaceutical industry is particularly fast-growing and highly competitive, so reducing time and costs associated to process optimization becomes instrumental to ensure speed to market and, consequently, profitability. The manufacturing of biopharmaceutical products, namely, monoclonal antibodies (mAbs), relies mostly on mammalian cell culture processes, which are highly dynamic and, consequently, difficult to optimize. In this context, there is currently an unmet need of analytical methods that can be integrated at-line in a bioreactor, for systematic monitoring and quantification of key metabolites and proteins. Microfluidic-based assays have been extensively and successfully applied in the field of molecular diagnostics; however, this technology remains largely unexplored for Process Analytical Technology (PAT), despite holding great potential for the at-line measurement of different analytes in bioreactor processes, combining low reagent/molecule consumption with assay sensitivity and rapid turnaround times.Here, the fabrication and handling of a microfluidic cartridge for protein quantification using bead-based affinity assays is described. The device allows geometrical multiplexed immunodetection of specific protein analytes directly from bioreactor samples within 2.5 h and minimal hands-on time. As a proof-of-concept, quantification of Chinese hamster ovary (CHO) host cell proteins (HCP) as key impurities, IgG as product of interest, and lactate dehydrogenase (LDH) as cell viability marker was demonstrated with limits of detection (LoD) in the low ng/mL range. Negligible matrix interference and no cross-reactivity between the different immunoassays on chip were found. The results highlight the potential of the miniaturized analytical method for PAT at reduced cost and complexity in comparison with sophisticated instruments that are currently the state-of-the-art in this context.