Advanced Materials Technologies

Bioxolography enables high-resolution fabrication of geometrically complex, cell-laden constructs for tissue engineering. However, tissue-relevant cell densities conflict with the optical transparency required for efficient dual-color volumetric printing. In this work, we extend the Bioxolography toolbox to include refractive index (RI) matching for cell-laden bioresins using iodixanol (IDX). Remarkably, IDX enhances optical transparency and boosts reactivity -- a phenomenon unique to Xolography. Yet, excessive IDX compromises dual-color efficiency through increased absorption and undesired UV-only curing, underscoring a central trade-off between optical clarity and photochemical performance. Systematic tuning of resin compositions along an iso-refractive index line demonstrated the versatility of Bioxolography, with IDX enhancing polymerization and 4-Hydroxy-TEMPO providing biocompatible inhibition. Optimizing composition and printing parameters yielded GelMA hydrogels with cell densities up to 5{middle dot}106 cells{middle dot}mL-1. Cell-laden prints achieved sub-100 {micro}m resolution and complex geometries such as channels and gyroids. Using skeletal muscle tissue as a model, we validated RI matched Bioxolography as a promising strategy for tissue engineering by demonstrating cell alignment along printed grooves and formation of mature muscle fibers characterized by MyoHC+ staining and fusion index. By integrating physical, chemical, and biological perspectives, this work advances Xolography toward biomaterials development and reinforces its position as an emerging volumetric (bio)printing technology. Table of Contents O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=175 SRC="FIGDIR/small/729865v1_ufig1.gif" ALT="Figure 1"> View larger version (86K): org.highwire.dtl.DTLVardef@407d86org.highwire.dtl.DTLVardef@1f116b4org.highwire.dtl.DTLVardef@109a437org.highwire.dtl.DTLVardef@1fc4ece_HPS_FORMAT_FIGEXP M_FIG C_FIG For printing higher cell density bioresins with Xolography, iodixanol (IDX) is added for refractive index-matching. The addition leads to an unexpected additional effect with increased reactivity in the dual-color photopolymerization. With careful adjustment of the resin composition and the printing parameters, Bioxolography is proven as a viable tool for tissue engineering.

Paper-based diagnostics such as lateral flow assays (LFAs) and microfluidic paper-based analytical devices ({micro}PADs) have attracted considerable attention because of their low cost, portability, and ease of use. Currently, to enable fabrication of {micro}PADs and improve LFA performance, hydrophobic blocks are patterned on paper substrates. However, fabrication of high-resolution hydrophobic barriers remains a major challenge. In this work, we developed a novel silicone extrudable ink for the fabrication of hydrophobic features on paper substrates. The ink was formulated using a vinyl-terminated polydimethylsiloxane (vPDMS) and polymethylhydrosiloxane (PMHS) system crosslinked through platinum-catalyzed hydrosilylation, and its rheological properties were tailored by incorporating silica fillers, obtaining a shear-thinning gel suitable for extrusion. The resulting formulation provided tunable properties, controlled deposition, and stable feature formation, enabling simple, low-cost, rapid, and robust fabrication of high-resolution hydrophobic barriers. Using this approach, we demonstrated improved fluid confinement and pattern fidelity on paper substrates, fabricated high-resolution paper microfluidic devices down to 150 {micro}m channel width, and enhanced the sensitivity of an LFA for a malaria diagnostic test. These results highlight the potential of this silicone ink platform as a practical and scalable strategy for advancing high-performance paper-based diagnostic technologies.

Oxygen availability is a critical yet all-too-often overlooked variable in organ-on-a-chip (OoC) systems. PDMS-based microfluidics remain the most common approach to facilitating oxygen equilibration with the incubator environment, but the materials tendency to ad- and absorb small hydrophobic molecules can pose significant concerns for pharmacological and toxicological studies. Yet there remains a lack of alternative gas-exchange materials feasible for OoC integration, even as the use of thermoplastic microfluidics in particular has otherwise proliferated. Here, we present commercially available track-etched nanoporous polycarbonate (50 nm pores, 1.18% porosity, [~]0.1 {euro}/cm2) as a practical alternative to polydimethylsiloxane (PDMS) for gas exchange in OoC. We show that nanoporous polycarbonate provides a thermoplastic material with an oxygen permeability of 3290 {+/-} 240 fs mol / kg, over an order of magnitude higher than PDMS. We demonstrate integration into existing lamination-based thermoplastic microfluidic fabrication workflows with sustained leak-free operation well above physiologically relevant pressures. We find that nanoporous polycarbonate does not compromise cell viability, but that high water vapor permeance necessitates a high-humidity environment around the device - though thickness-normalized water vapor permeability is notably similar to PDMS. We validate the OoC application with Caco-2 intestinal epithelial cells by monitoring oxygen levels during the critical cell attachment phase, with nanoporous polycarbonate allowing for maintenance of stable oxygen tension, in stark contrast to severe hypoxia in nonporous controls within 30 minutes. We further show that this uncontrolled hypoxia correlates with a time-delayed increase in cellular hypoxia inducible factor-1 reporter expression. Overall, our findings position nanoporous polycarbonate as a low-cost, mechanically robust, and fabrication-friendly alternative that can bring controlled oxygen availability to PDMS-free microfluidics and OoC.

Over the past decade, the integration of microgel-based granular hydrogels in biomedical technologies has experienced substantial growth due to the numerous benefits microgels offer. However, the inability to easily adopt uniform microgel fabrication workflows at scale constitutes a major bottleneck, or in some cases, a barrier-to-entry that stunts further growth of the field. The gold-standard technique for emulsion-based microgel production is through microfluidic droplet-generating devices that produce liquid gel precursor droplets that gel post-production. However, traditional microfluidic workflows often require multiple independent flows and controlled pressure sources, along with a steep learning curve in using microfluidics to achieve uniform droplet sizes reproducibly and repeatedly. This difficulty in adopting microgel fabrication is further compounded by low throughput and the extensive flow rate calibration required when switching to new formulations (e.g., material type, droplet size). In this work, we present a step-emulsion system that bridges the gap by providing a robust and simple setup. We experimentally characterize and evaluate how flow and outlet channel dimension contribute to the generation of uniform droplet populations at specific sizes. With our large dataset consisting of various outlet channel dimensions, we evaluated outlet channel geometrical impacts (height, width, cross-sectional area, aspect-ratio, etc.) on gel precursor droplet size and generation throughput. We demonstrate robust, highly compatible, and repeatably uniform droplet generation from various gel precursor polymer backbones, users with varying microfluidics experience, and a wide viscosity range, including alginate solutions with 650 times the viscosity of water. Furthermore, we confirmed consistent gel precursor droplet generation outcomes driven by a constant flow source (syringe pump) and by direct manual injection as a simple and highly adoptable option for the generation of gel precursor droplets. This platform is ideal for researchers seeking rapid and easy microgel fabrication, regardless of microfluidics experience.

Organ-on-chip (OoC) systems enable the recapitulation of key structural and functional characteristics of human tissues within controlled micro-engineered environments. In mechanically active tissues such as musculoskeletal, cardiac, and vascular systems, the incorporation of dynamic physical forces is essential for replicating the biomechanical cues governing cellular morphology and functional responses in-vivo. Without such stimuli, OoC models may fail to capture physiologically relevant tissue behaviors. Porous and semi-permeable membranes are critical components of OoCs, facilitating selective transport of nutrients, gases, and signaling molecules between cellular compartments to support biologically accurate barrier replication. Hence, fabrication strategies that permit precise modulation of membrane permeability are desirable to accommodate for the varying needs in pore size and porosity across organ systems. This study presents a two-stage fabrication process for stretchable, microporous polydimethylsiloxane (PDMS) membranes using femtosecond (fs-) pulse laser drilling. The laser-ablated pores exhibit a characteristic conical morphology, with diameters tapering from the laser entry to exit point. By modulating laser power and number of pulses, 6-15 m exit-end pore diameters were achieved in 50 m thick PDMS films. The membranes demonstrated strong mechanical resilience, with a 5-12% reduction in Youngs modulus after 500 cycles of strain loading. Furthermore, membranes fabricated at lower laser powers exhibited superior retention of elasticity, highlighting the influence of processing parameters on mechanical behavior. Cytocompatibility and permeability assessments confirmed that the membranes supported sustained cell viability and proliferation over at least three days. In size-restricted membrane pore geometries, cellular migration was constrained without any inhibition of biomolecular transport. This selective permeability is critical in multilayer OoC architectures, where a balance between biomolecular diffusion and cellular compartmentalization is necessary to preserve distinct tissue interfaces and functional organization. This work presents fs-laser micro-drilling as a robust and tunable fabrication strategy for producing mechanically resilient, selectively permeable PDMS membranes for physiologically relevant OoC applications.

Conventional extrusion-based 3D bioprinting encounters limitations in fabricating intricate tissue architectures due to fixed nozzle diameters and fixed deposition orientations. These constraints restrict conformal printing on curved or non-planar surfaces and often necessitate support-intensive fabrication strategies. This work introduces a mechanically simplified extrusion platform inspired by the swivel jet nozzle, featuring a free-degree-of-orientation extrusion head termed the universal extrusion head (Univ-Ex head), coupled with a modular nozzle architecture. The Univ-Ex head employs a swivel-like mechanical design that enables orientation freedom without external actuation in its current implementation, thereby minimizing mechanical complexity while supporting deposition on physiologically relevant, non-planar geometries. Multiple nozzle concepts were developed through comparative CAD iterations, with two representative geometries--a flat nozzle and a conical nozzle--selected for experimental validation. The platform is evaluated through parametric CAD design, stereolithography-printed prototypes, proof-of-concept extrusion experiments, and fluid dynamics simulations performed using FLOW-3D software. Numerical and experimental results demonstrate stable filament formation and clear diameter-dependent extrusion behavior, while simulations further confirm the feasibility of angled and non-planar deposition. A variable-diameter nozzle concept is proposed as a forward design direction to enable real-time adjustment of bioink flow rate and deposition resolution in principle; however, the present study intentionally validates the system using fixed-diameter nozzle variants to maintain stable numerical and experimental boundary conditions. A gear-integrated Univ-Ex head is also presented as a forward upgrade and demonstrated as a single-piece prototype. Collectively, this work establishes a scalable, hardware-focused pathway toward conformal bio-additive manufacturing. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=96 SRC="FIGDIR/small/734010v1_ufig1.gif" ALT="Figure 1"> View larger version (63K): org.highwire.dtl.DTLVardef@8833caorg.highwire.dtl.DTLVardef@33dforg.highwire.dtl.DTLVardef@14d8d11org.highwire.dtl.DTLVardef@685ef0_HPS_FORMAT_FIGEXP M_FIG C_FIG

Reliable and scalable soft implantable neural interface fabrication remains a key challenge for chronic bioelectronic applications. Here, we present a transparent soft microelectrode fabricated with electrohydrodynamic (EHD) printing, utilizing the fluorinated polymer poly(vinylidene fluoride-co-hexafluoropropylene) (PVDF-HFP) and poly (3, 4-ethylenedioxythiophene) polystyrene sulfonate (PEDOT: PSS) to form seamless, selectively patterned multilayer structures with low impedance and long-term stability. Controlled in situ curing during printing yields dense, void-free substrate and encapsulation layers, suppressing interfacial defects and ionic pathways, while maintaining high optical transparency (>60%) with PEDOT:PSS. The printed microelectrodes exhibit low impedance, high charge storage and injection capacities, and stable electrochemical behavior under biomimetic conditions. In addition, the devices demonstrate robust mechanical and electromechanical stability under cyclic deformation in both dry and wet environments, as well as under prolonged electrical stimulation. Accelerated aging studies project multi-year operational lifetimes, and in vitro/in vivo biocompatibility assessments confirm excellent tissue integration. These results establish EHD-printed fluorinated polymer-based microelectrodes as a scalable and durable platform for chronic implantable biointerfaces. ToC O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=182 SRC="FIGDIR/small/726391v1_ufig1.gif" ALT="Figure 1"> View larger version (79K): org.highwire.dtl.DTLVardef@152c58aorg.highwire.dtl.DTLVardef@126f1f5org.highwire.dtl.DTLVardef@1d743cforg.highwire.dtl.DTLVardef@1a4d743_HPS_FORMAT_FIGEXP M_FIG C_FIG This report presents an electrohydrodynamically printed transparent soft microelectrode for chronic purposes. Electrohydrodynamic printing promotes seamless multilayer structures with selective deposition and long-term mechanical stability. The devices show low impedance, high charge capacity, and robust electrochemical/electromechanical properties. Accelerated aging projects [~]7.2 year lifetimes, and XPS/SEM-EDS confirm strong ion barrier properties and biocompatibility for chronic implantation.

Traditional bioelectronic devices are limited by poor biointerfacing due to their substantial mismatch in mechanical and biochemical properties. In tissue engineering, soft and bioactive materials support biointegration by harnessing or mimicking the natural extracellular matrix (ECM). Building bioelectronic devices from ECM should improve their biointegration, yet there are limited methods to fabricate them due to current manufacturing approaches. An additive manufacturing strategy is presented here for collagen-based bioelectronic interfaces that integrates conducting polymer electrodes with ECM-based substrates or encapsulation layers. Addition of poly(ethylene glycol) diglycidyl ether (PEGDE) to poly(3,4-ethylenedioxythiophene):poly(styrene sulfonate) (PEDOT:PSS) colloidal dispersions enables direct extrusion-based patterning under mild conditions compatible with collagen substrates, and forms aqueous stable and highly conducting printed patterns (2788 S m-{superscript 1}). The resulting interfaces maintain stable electrochemical performance over 7 days in physiological environments, and support primary human cell adhesion, viability, and proliferation across both material regions. A sacrificial patterning strategy using 3D printed cacao butter further enables spatial control of collagen encapsulation. This approach establishes a framework for fabricating functional bioelectronic devices based on ECM to further enhance device biointerfaces for tissue models and implantable systems.

Foreign body response is a common yet serious challenge for biomedical implants. It can trigger inflammation and eventually lead to the formation of a fibrotic capsule, which compromises device function. Although significant efforts have been made to develop antifibrotic surface coatings for implantable materials, developing broadly applicable solutions remains challenging due to the diversity of materials used in biomedical implants. Here, we propose a simple and versatile strategy to develop antifibrotic coatings for biomedical implants. Photoreactive benzophenone groups are incorporated into designer polymers to enable covalent attachment to various substrates. The effect of benzophenone group density within polymer chains on surface coating efficiency was investigated, and an optimal BP incorporation ratio was identified. Polymers incorporating varying ratios of an anti-fibrotic small molecule and anti-fouling zwitterionic moieties were synthesized and successfully attached to silicone implants. In vivo evaluation of these implants in C57BL/6 mice identified an optimized polymer composition that reduced fibrotic capsule thickness by around 60%. Coating of commercial medical catheters with this optimized polymer reduced collagen deposition by over 3.5-fold following 4 weeks of implantation in the peritoneal space of C57BL/6 mice. Finally, we demonstrated that the optimized polymer coating can be readily applied to a variety of commonly used biomedical materials using this straightforward method, highlighting the versatility of the approach. This work provides a facile and broadly applicable strategy for developing antifibrotic coatings, which has the potential to expand the design of surface modifications aimed at improving the performance of biomedical implants.

Mechanical forces have a major effect on cell behavior. Most cells in vitro are grown under static conditions on hard tissue culture plastic, conditions that do not accurately reflect living tissues. The ability of cells to sense and respond to mechanical forces is essential for key biological processes, including development, proliferation, and migration. Disruption of the ability to respond to mechanical forces are known to be a critical factor in many diseases, including cardiovascular disease, progeria, and cancer. Here, we present the design, fabrication, and biological testing of a custom-built cell-stretching device that applies controlled biaxial strain to cells cultured on a polydimethylsiloxane (PDMS) membrane. We then used this device to examine how cells respond to strain. In response to biaxial strain, MCF-7 cells activated the mechanosensitive immediate early gene (IEX-1), with its expression increasing significantly after 1 and 3 hours of stretching. Cells exposed to mechanical strain also remodeled their cytoskeleton in a direction-dependent manner. Under uniaxial strain, actin filaments reoriented perpendicular to the stretch direction, whereas biaxially stretched cells do not promote directional reorientation, but instead appear to reinforce actin at the cell periphery. Similarly, cells under uniaxial strain exhibited changes in nuclear orientation and shape that were not observed under biaxial strain. Nuclear area remained unchanged in either strain condition. These results highlight that the biaxial stretcher can be used to apply strain to cells, and that cells respond differently to biaxial strain compared to what has been reported for uniaxial strain.

Cancer is a significant contributor to global mortality and places a substantial burden on healthcare systems, underscoring the need for improved strategies for developing and evaluating new therapies. Electrochemical impedance monitoring of in vitro cancer models is a promising technique for evaluating treatment effectiveness, particularly for evaluating how well a drug may kill cancer cells. This approach is advantageous over conventional end-point assays because it is non-destructive, label-free, and can provide temporal information on cell behavior and drug kinetics. However, traditional impedance devices are limited in that they do not support three-dimensional cell culture that has become standard in cancer studies. Typical devices are planar substrates that support monolayer culture, which has been shown to overestimate drug effectiveness. In this work, we propose 3D printed bioelectronic scaffold devices that provide 3D cancer cell culture while functioning as an on-chip readout for monitoring changes in cell characteristics via impedance. We describe device development and demonstrate reproducible fabrication, stable electrochemical properties, cell detection by impedance, and proof-of-concept monitoring of cytotoxicity in response to a chemotherapeutic drug. Overall, this technology offers a promising platform that could be further developed for compound screening as part of drug development or precision medicine.

Resin three-dimensional (3D) printing is an increasingly popular manufacturing and prototyping method used to create microphysiological systems (MPS), but resin cytotoxicity significantly hinders its adoption, especially when sensitive cell models are incorporated. The mammalian oocyte and early preimplantation embryo consist of cells that are highly sensitive to toxicants and thus represent stringent cell-based models for biocompatibility testing. We developed a Multi-Endpoint Oocyte Safety Assay (MEIOSA) to evaluate the biocompatibility of four ISO 10993 biocompatible BioMed resins (Clear, Durable, Elastic 50A, and Flex 80A). MEIOSA assesses the viability, morphology, meiotic stage, and meiotic spindle morphology of the oocyte after in vitro maturation (IVM). Oocytes were in vitro matured in plate inserts 3D printed with the four BioMed resins. Oocytes cultured in rigid resins (Clear and Durable) or elastomeric resins (Elastic 50A, and Flex 80A) exhibited impaired meiotic progression and complete oocyte degeneration, respectively, relative to controls cultured in polystyrene which matured normally. To determine whether such cytotoxicity could be prevented, we coated the resin inserts with a 5 {micro}m impermeable Parylene-C (PC) barrier. PC coating completely rescued the degeneration and meiotic maturation defect phenotypes for all resins. Remarkably, when the most cytotoxic material (Flex 80A) was coated with PC, the resulting eggs were fertilization-competent and produced embryos capable of normal preimplantation development via in vitro fertilization. Our findings demonstrate that standardized viability-based biocompatibility tests do not identify cytotoxic effects for all cell types and establish MEIOSA as a high sensitivity test model to robustly evaluate biomaterial biocompatibility. Furthermore, PC coating prevents the toxic effects of all resin-3D-printed materials tested, opening up a new toolbox to create MPS compatible with reproductive, and by extension, other sensitive cell cultures. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=197 HEIGHT=200 SRC="FIGDIR/small/730268v1_ufig1.gif" ALT="Figure 1"> View larger version (71K): org.highwire.dtl.DTLVardef@18fab44org.highwire.dtl.DTLVardef@1352e5forg.highwire.dtl.DTLVardef@77889forg.highwire.dtl.DTLVardef@1aabd89_HPS_FORMAT_FIGEXP M_FIG C_FIG

Engineered epithelial models require three-dimensional extracellular matrix environments that support organized cell growth and allow independent access to luminal and basal compartments. However, many organ-on-chip (OoC) fabrication strategies rely on planar geometries, non-native materials, or multi-step assembly workflows that limit architectural complexity and experimental control. Here, we report a direct in-chip volumetric printing strategy for fabricating stretchable and perfusable collagen-I scaffolds inside custom OoC devices. A vitamin C-regulated ruthenium/sodium persulfate photocrosslinking system enabled high-fidelity printing of collagen-I into open-lumen architectures with ductal- and alveolar-inspired features. By generating scaffolds directly within the final culture device, this workflow eliminates post-print transfer and integrates defined collagen architectures with compartmentalized fluidic access and a mechanically actuable chip format. To support chip-based culture, printed collagen constructs were stabilized after fabrication using EDC/NHS chemistry, which limited thermally induced collagen densification, improved shape retention, and maintained scaffold anchorage during perfusion. The chip design provided separate access to the printed lumen and surrounding basal compartment, which enabled compartment-specific fluid handling while preserving scaffold integrity during inflation, stretching, and perfusion of the printed construct. On the collagen-I scaffolds, human milk-derived mammary epithelial cells formed epithelial layers with tight junctions and lactation associated markers. The platform further supported perfusion culture, in situ staining, and whole-chip volumetric imaging. Together, this work establishes direct in-chip collagen-I volumetric printing as a biofabrication strategy for creating perfusable epithelial tissue chips with native matrix architecture and compartmentalized fluidic control.

Sensitive and low-cost protein biomarker detection is critical for disease diagnosis. Advanced microfluidic systems can generate miniature reaction compartments for a high-sensitivity assay. However, these platforms often require external instruments, skilled operators, and complex setups. Here, we develop a Lab on a Capillary (LabCap) platform that integrates photopatterned hydrogel rings within a glass capillary using a reconfigurable stop-flow lithography system. During sample loading and unloading steps, nanoliter-scale aqueous droplets (torodrops) are spontaneously formed around the hydrogel rings, creating isolated reaction compartments without the need for external instruments or an immiscible oil phase. The LabCap platform enables quantitative detection of clinically relevant biomarkers, including C-reactive protein (CRP) and N-terminal pro-B-type natriuretic peptide (NT-proBNP). By adjusting the incubation protocol, assay speed and sensitivity can be tuned to meet different analytical requirements. A periodic medium exchange protocol enables biomarker detection at concentrations as low as 1 ng/mL, whereas prolonged static incubation extends detection to 0.1 ng/mL. In addition, LabCap offers practical advantages, including low fabrication cost (< EUR 1 per device), low reagent consumption (<100 microlitres per assay step), and minimal wash-buffer usage (1 mL). These results demonstrate that LabCap is a simple, cost-effective, and versatile platform for biomarker detection.

This work presents a facile and rapid fabrication method for high-performance bioelectrode coatings through electropolymerization of poly(3,4-ethylenedioxythiophene) (PEDOT) and polydopamine (PDA). Expanding on previous work, we develop a cyclic voltammetry electropolymerization process to deposit PEDOT:PDA coatings on 2 mm gold electrode substrates. The coatings exhibit a 40-fold increase in cathodic charge storage capacity ([~] 40mC{middle dot}cm-2) and significant impedance modulus reduction compared to uncoated gold electrodes. Morphological characterization revealed a uniformly porous surface that corroborates the enhancement in electrochemical performance. Our scalable approach offers a promising option to fabricate bioelectrodes for application to neural interfaces and implantable and wearable bioelectronics. Clinical RelevanceThe improved electrochemical performance and scalable fabrication of PEDOT:PDA coatings support their potential to enhance the stability and signal quality of neural and implantable bioelectronic devices.

3D spheroids, which closely replicate three-dimensional cell-cell and cell-extracellular matrix interactions, offer superior predictive capabilities compared to conventional 2D monolayer cultures, positioning them as forward-looking platforms in drug testing, cancer biology, and regenerative medicine. However, high-throughput generation of uniform sized spheroids is still a technological challenge. In one hand, the use of conventional ultra-low attachment (ULA) multiwell plates for this purpose is labour intensive and complex. On the other hand, the use of microfabricated facilities demands cutting-edge infrastructure such as clean room, photolithography, and microfluidic setup which are often unavailable for the resource constrained laboratories. In this study, we addressed these problems by developing a low-cost Do-It-Yourself (DIY), polydimethylsiloxane (PDMS) and agarose-based spheroid generation device, capable of producing and maintaining hundreds of spheroids with minimal user intervention. We have demonstrated two variants based on their size, termed here as S1 and S2 devices which fit into 6-well and 12-well plates, and can generate 600 and 1200 uniform-sized spheroids respectively. We validated our device with various cell lines including primary and cancerous cell lines. We further demonstrated the drug testing capabilities of the device by estimating the IC50 value of the anticancer drug Temozolomide on U87-MG. The value was comparable with the same obtained from the spheroids generated using conventional ULA plates. Additional attachment of a perfusion system made the device suitable for long-term spheroid culture without much user intervention. Furthermore, the devices can also be used for the production of spheroids with gradually changing diameters in a controlled manner, resembling a size gradient. This feature is useful for checking the effect of drugs on different-sized spheroids and for co-culturing spheroids with varying cell densities, mimicking the disease architecture. We have co-cultured two types of the placental trophoblast cells, i.e., extravillous trophoblast (HTR-8) and syncytiotrophoblast (BeWo) with varying densities. In summary, this paper demonstrates a unique DIY method for a high-throughput uniform-sized spheroid generation at a fraction of cost which can be deployed to resource-constrained labs.

Automation of organoid and cell culture processes is essential for achieving scalable and standardized experimentation in regenerative medicine and stem cell research. However, existing microfluidic platforms often rely on complex setups, limiting their integration within standard incubator environments. To address these challenges, we developed a compact, scalable multi-well platform featuring 3D-printed, servo-actuated disposable microvalves for fully automated media and drug exchange. This design eliminates the need for external pressure sources and control channels, providing a simplified and cost-effective solution for organoid culture. The platform integrates an internet-connected microscopy module with a motorized XYZ stage, allowing continuous, real-time imaging of individual wells directly within the incubator. It supports precise and reliable fluid handling under physiological conditions, improving throughput, reproducibility, and accessibility. We validate the platform through bench-top testing and in both mouse and human organoid models. Morphological analysis, immunohistochemistry (IHC), and qPCR demonstrate comparable viability, growth, and gene expression profiles between automated and manual culture conditions. These results establish a robust and scalable framework for fully automated organoid culture, offering a simplified and accessible alternative to conventional microfluidic systems with broad applications in regenerative medicine, drug discovery, and scalable biological screening. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=101 SRC="FIGDIR/small/732526v1_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@5efd07org.highwire.dtl.DTLVardef@3600d0org.highwire.dtl.DTLVardef@16f85f5org.highwire.dtl.DTLVardef@c39fbd_HPS_FORMAT_FIGEXP M_FIG C_FIG

The combination of synthetic biology and additive manufacturing has driven major changes in production of biomaterials, especially through the use of three-dimensional (3D) bioprinting to create engineered living materials. However, current fabrication methods can be limited by prohibitive hardware costs and the inability to maintain structural fidelity in complex, free-form living architectures. This work demonstrates how to build a low-cost, open-source 3D bioprinting platform that can make complicated bacterial structures with complex geometry and high dimensional accuracy. A commercially available, conventional fused deposition modeling 3D printer was modified to create a bioprinting system that is simple to build. The modified bioprinter, which costs around $450, is less expensive than many commercial bioprinters. This 3D-printing technology uses slurry-based support bath methods featuring low-cost gelatin and agarose microparticles, resulting in structures with a high aspect ratio (>8:1) and feature sizes as small as 260 m. The optimization of critical printing settings, including the ability of the bioink to retract during non-print movements, resulted in a reduction of unwanted bacterial deposition by nearly two orders of magnitude. Long-term viability experiments showed that bacteria in the bioprints could survive for at least 28 days with nutrient supplementation. Additionally, 3D-printed engineered biofilms revealed that incubation conditions and extracellular matrix composition significantly impacted the mechanical properties of printed constructs, with tradeoffs between matrix production and mechanical integrity. This study showcases an accessible 3D bioprinting platform for advanced bioprinting technologies, enabling development of engineered living materials with potential applications in synthetic biology, biotechnology, and tissue engineering.

Background and ObjectivesCardiovascular cells differentiated from human induced pluripotent stem cells (iPSCs), including cardiomyocytes, are valuable for evaluating human cardiac pharmacology and toxicity. Early assessment of cardiotoxicity, especially for novel drugs like anticancer agents, is essential for improving drug development efficiency and reducing costs. This study aimed to develop a highly sensitive bioassay system capable of evaluating the physiological function of human cardiac tissue in vitro. MethodsHuman iPSCs were differentiated into cardiovascular cell types (cardiomyocytes, vascular endothelial cells, and vascular mural cells) and assembled into a cardiac tissue model on aligned fiber device. This tissue was cultured dynamically to induce the formation of vascular network-like structure. By combining the fiber device with our previously developed heart-on-a-chip microdevice (HMD), we created a new model of HMD (Aligned Fiber-based HMD; AF-HMD) with improved throughput and stability. Pulsatile force changes induced by drug exposure were quantified by tracking the displacement of fluorescent microbeads within the microchannels. ResultsAF-HMD demonstrated functional responses to known cardiac agonists and toxicants, such as doxorubicin. The device also replicated clinically relevant cardiotoxic events, including the synergistic effects of trastuzumab and doxorubicin, showing marked reductions in contractile force and beat rate, mirroring clinical observations. ConclusionsThe AF-HMD system provides a sensitive and reproducible platform for evaluating cardiotoxicity in drug development. It offers a promising tool for preclinical screening, with potential applications in personalized medicine and predicting cardiotoxic risk in cancer therapy.

Optical fiber-assisted printing (OFAP) was recently introduced as a straightforward light-based platform for the spatially controlled photopolymerization of hydrogel-based resins. Here, we extend this concept toward optical fiber-assisted bioprinting (OFAB) by processing cell-laden GelMA- and GelMA/PEGDA-based bioresins in a freeform embedded printing configuration. The system relies on a 405 nm LED-coupled optical fiber mounted on an automated 3D motion platform, enabling localized photocrosslinking directly within a resin bath. First, GelMA and GelMA/PEGDA formulations containing LAP and tartrazine were screened to evaluate the influence of light intensity, printing velocity, and material composition at the line width and curing depth. Single-line features with widths down to 70 {+/-} 20 {micro}m were obtained under optimized conditions, while more robust printing conditions yielded reproducible features in the range of 200-300 {micro}m. Photorheological and rotational rheology measurements confirmed that the formulations provide both thermoresponsive support during printing and photocrosslinked stability after processing. The incorporation of L929 cells demonstrated high cytocompatibility for GelMA and GelMA/PEGDA 6000 Da formulations, with viabilities above 90% after 7 days for selected printed constructs. Importantly, increasing the cell concentration up to 1 x 107 cells mL-1 did not prevent printing and reduced the extent of overcuring, suggesting that cell-induced turbidity can improve spatial confinement of polymerization in OFAB. Finally, a customized OFAB printer was developed to enable temperature-controlled processing and the fabrication of centimeter-scale 3D structures, including cell-laden constructs. Overall, this work establishes OFAB as an accessible and modular bioprinting strategy for cell-laden and optically turbid hydrogel resins, complementing existing light-based biofabrication approaches.