Culture Device Research Articles

A novel layered culture device reveals spatial dynamics of root element uptake and optimal silicon application site for mitigating chromium uptake by rice

Understanding root uptake mechanisms for various elements is crucial for optimizing heavy metal remediation strategies and enhancing plant-nutrient interactions. However, simple and effective methods to differentiate the contributions of specific root segments in element uptake are lacking. Here, we developed a layered culture device consisting of a culture box and a plant suspension mechanism, which isolates different root segments through solid media and waterproof coating. Then, we used the device to investigate the roles of distinct root segments (0–1 cm and 1–2 cm from the tip) in heavy metal chromium (Cr) and beneficial element silicon (Si) uptake in rice. The results indicated that the 0–1 cm root segment contributed approximately 58% of leaf Cr(VI), with higher efflux compared to the 1–2 cm segment. Conversely, the 1–2 cm root segment served as the primary source of leaf Si and Cr(III), accounting for 62% and 54%, respectively. The translocation factors for Cr(VI) were similar for both segments (0.039 and 0.032), while the Cr(III) translocation factor for the 0–1 cm root segment (0.061) was 2.8 times that of the 1–2 cm segment. Notably, Si application to the 0–1 cm segment most effectively alleviated Cr (III) and Cr (VI) stress, boosting shoot length, fresh weight, and chlorophyll concentration and reducing Cr concentrations in roots and leaves by 24.7%–65.7%. In contrast, Si application to the 1–2 cm segment had minimal impact on rice growth or Cr uptake. These results suggest a deep Si application strategy for remediating Cr-contaminated soil. The innovative device provides a scientific foundation for distinguishing element uptake contributions of different root segments and enhancing the utilization efficiency of remediation materials and nutrient management in agriculture.

Rotating culture regulates the formation of HepaRG-derived liver organoids via YAP translocation

BackgroundLiver organoid serves as an alternative model for liver pathophysiology in carbohydrate or lipid metabolism and xenobiotic metabolism transformation. Biomechanical cues including spaceflight mission can affect liver organoid construction and their related functions, but their underlying mechanisms are not fully understood yet. Here, a rotating cell culture device, namely Rotating Flat Chamber (RFC), was specifically designed for adhering cells or cell aggregated to elucidate the effects of altered gravity vector on HepaRG-derived liver organoids construction.ResultsThe organoids so formed under RFC presented the fast growth rate and large projection area. Meanwhile, the expressions of two pluripotency markers of SOX9 and CD44 were enhanced. This finding was positively correlated with the increased YAP expression and nuclear translocation as well as the elevated α4β6-integrin expression. Inhibition of YAP expression and nuclear translocation decreased the expression of SOX9 and CD44 under RFC, thereby attenuating the pluripotency of HepaRG-derived liver organoids.ConclusionsIn conclusion, we proposed a novel liver organoid construction method using rotating culture, by which the pluripotency of liver organoids so constructed is mediated by α4β6-integrin and YAP translocation. This work furthered the understanding in how the gravity vector orientation affects the construction of liver organoids and the related mechanotransductive pathways.

To evaluate and compare the effect of diabetes and complete denture wearing on the occurrence of Mucor isolates in edentulous geriatric patients

Objectives: The objective of this study was to explore the impact of diabetes and complete denture use on the presence of Mucor isolates in edentulous geriatric patients. Material and Methods: Intraoral swab samples were collected using a sterile culture device, focusing on the center of the hard palate for all patient groups. Immediately after collection, each sample was immersed in a sterile culture-collecting device containing normal saline to prevent swab drying. Subsequently, these swabs were isolated and cultured in Sabouraud’s dextrose agar medium at a temperature range of 25–30°C. Results: Statistical analysis using the Chi-square test revealed no Mucor isolates in any of the four groups. However, the combination of diabetes and complete denture use led to the isolation of other commensal fungi in 65% of samples, contrasting with the non-diabetic, non-denture-wearing group where this occurrence was only 5%. Conclusion: The study concluded that there was no observable correlation between the occurrence of Mucor isolates in edentulous geriatric patients, irrespective of complete denture use, or diabetic status. Nevertheless, the coexistence of diabetes and complete denture use significantly increased the presence of opportunistic fungi in the oral cavity compared to non-diabetic, non-denture-wearing patients.

Combining fermentation broth with spent water recycling for microalgae cultivation: Nutrient reutilization, biodiesel production and techno-economic simulation

This study presents an innovative and promising technology, Water Cycling Combined Fermentation Broth (WC2FB), which offers a sustainable solution to significantly reduce the costs associated with microalgae-based biodiesel production. The innovative approach involves utilizing food waste as a nutrient source, supplemented by recycled water after microalgae harvesting, with the goal of improving the viability of industrial biofuel production. The results of nutrient removal efficiency demonstrated that the addition of 15% fermentation broth to the microalgal suspension efficiently utilized volatile fatty acids, resulting in a significant increase in lipid content. Additionally, biocomponent analysis confirmed that the WC2FB mode did not compromise the desirable properties of biodiesel. Through techno-economic simulations, we predicted a 90% probability of achieving net present value (NPV) at a selling price of $5/kg. Finally, we recommended that directing future efforts be directly towards the development of culture devices, low-cost bioflocculation methods, and biorefinery extraction to further advance biodiesel production.

Shrinking Cancer Research Barriers: Crafting Accessible Tumor‐on‐Chip Device for Gene Silencing Assays

Tumor‐on‐chip (ToC) is crucial to bridge the gap between traditional cell culture experiments and in vivo models, allowing to recreate an in vivo‐like microenvironment in cancer research. ToC use microfluidics to provide fine‐tune control over environmental factors, high‐throughput screening, and reduce requirements of samples and reagents. However, creating these microfluidic devices requires skilled researchers and dedicated manufacturing equipment, making widespread adoption cumbersome and difficult. To address some bottlenecks and improve accessibility to ToC technology, innovative materials and fabrication processes are required. Polystyrene (PS) is a promising material for microfluidics due to its biocompatibility, affordability, and optical transparency. Herein, a fabrication process based on direct laser writing on thermosensitive PS, allowing the swift and economical crafting of devices with easy pattern alterations, is presented. For the first time, a device for cell culture fabricated only by PS is presented, allowing customizing and optimization for efficient cell culture approaches. These biochips support 2D and 3D cultures with comparable viability and proliferation kinetics to traditional 96‐well plates. The data show that gene and protein silencing efficiencies remain consistent across both chip and plate‐based cultures, either 2D culture or 3D spheroid format. Although simple, this approach might facilitate the use of customized chip‐based cancer models.

A laying on of hands: reading afrodiasporic spirituality, matrilineage & collectivism in lemonade

ABSTRACT Lemonade (2016) is a genre-blending text that imports cultural and fantastical devices in order to revise, interrogate, and re-examine history–specifically, the traumas inherited from enslavement. In its analysis of history, Lemonade immortalizes Afrodiasporic spiritual practices and enshrines Black women-healers and magic-users as folk heroes. This work advances that the Afrodiasporic spiritual practices and gynocentric connections are used to form Black female collectives. It draws largely on Black and Africana feminist and womanist scholarship to frame an expanded discussion of spirituality and Black motherhood in the American South, and to demonstrate how collectives–sacred, transhistorical communities–are critical to Black female healing. It contextualizes Lemonade’s “chapters,” visuals, and wider narrative, and it examines the Southern psychic and epistemological resources that Beyoncé, as protagonist, consults. In doing so, this work ultimately intends to create new intellectual space for readers to revisit questions of collectivity in Lemonade and consider the indispensability of community for Black women and girls.

3D-printed microfluidic and millifluidic devices for cell culture and mechanotransduction studies: A review

The practice of conventional cell culture techniques affects the quality of biological research in mechanobiology by not replicating the cell’s microenvironment adequately. Microfluidic devices resembling 3D in vivo cell culture offer viable alternatives for mechanobiology studies and other applications including cell screening, cell separation, and point-of-care diagnostics. This review highlights recent advances in additive manufacturing (AM) to fabricate microfluidic and millifluidic devices through fast design iterations and cost reduction compared with conventional device manufacturing. Key advances in AM technologies such as fused deposition modeling (FDM), stereolithography apparatus (SLA), and digital light processing (DLP) have allowed the direct manufacture of complex microfluidic devices with master molds and sacrificial structures using multiple material components. Currently, available 3D printed devices for the precise fabrication of milli- and microfluidic devices, essential to mimicking the cellular microenvironment, while economically reducing production costs by eliminating expensive clean-room environments and reducing material waste are highlighted.

Mechanistic study of a low-power bacterial maintenance state using high-throughput electrochemistry

Mechanistic studies of life's lower metabolic limits have been limited due to a paucity of tractable experimental systems. Here, we show that redox-cycling of phenazine-1-carboxamide (PCN) by Pseudomonas aeruginosa supports cellular maintenance in the absence of growth with a low mass-specific metabolic rate of 8.7×10-4 W (g C)-1 at 25°C. Leveraging a high-throughput electrochemical culturing device, we find that non-growing cells cycling PCN tolerate conventional antibiotics but are susceptible to those that target membrane components. Under these conditions, cells conserve energy via a noncanonical, facilitated fermentation that is dependent on acetate kinase and NADH dehydrogenases. Across PCN concentrations that limit cell survival, the cell-specific metabolic rate is constant, indicating the cells are operating near their bioenergetic limit. This quantitative platform opens the door to further mechanistic investigations of maintenance, a physiological state that underpins microbial survival in nature and disease.

Automated counting and classifying Daphnia magna using machine vision

Daphnia magna (D. magna) is a model organism widely used in aquatic ecotoxicology research due to its sensitivity to environmental changes. The survival and reproduction rates of D. magna are easily affected by toxic environments. However, their small size, fragility, and transparency, especially in neonate stages, make them challenging to count accurately. Traditionally, counting adult and neonate D. magna relies on manual separation and visual observation, which is not only tedious but also prone to inaccuracies. Previous attempts to aid counting with optical sensors have faced issues such as inducing stress damage due to vertical movement and an inability to distinguish between adults and neonates. With the advancement of deep learning technologies, our study employs a simple light source culture device and utilizes the Mask2Former model to analyze D. magna against the background. Additionally, the U-Net model is used for comparative analysis. We also applied OpenCV technology for automatic counting of adult and neonate D. magna. The model's results were compared against manual counting performed by experienced technicians. Our approach achieves an average relative accuracy of 99.72 % for adult D. magna and 98.30 % for neonate. This method not only enhances counting accuracy but also provides a fast and reliable technique for studying the survival and reproduction rates of D. magna as a model organism.

A single-cell 3D dynamic volume control system for chondrocytes.

In articular cartilage, zone-specific cellular morphology is a typical characteristic of cartilage tissue, which is related with chondrocyte function, inflammation and osteoarthritis (OA). Chondrocyte hypertrophic phenotype is a criticle physiological process which indicates a hallmark of chondrocyte terminal differentiation and bone formation. Thus, developing a in vitro cell culture system for dynamic regulation of single chondrocyte volume at a three-dimensional (3D) level is particularly necessary for understanding how physical cues of matrix microenvironment regulate chondrocyte fate and the degeneration of articular cartilage. Here, based on the soft lithography techniques, we have constructed well-defined single-cell 3D dynamic volume control system to recapitulate the physiological matrix microenvironment of single chondrocyte niche. The results of finite element analysis indicated that the stress and strain distribution in the cell culture region is homogeneous during the stretching process. Additionally, 3D dynamic volume expansion and compression of single cells in physiological or hyperphysiological can be realized in this cell culture system. Our device for single-cell 3D dynamic culture provides a microphysiological culture system for chondrocytes to explore the mechanisms of cartilage hypertrophy, as well as develops a new paradigm for functional cartilage tissue engineering and regenerative medicine.

Development of specialized devices for microbial experimental evolution.

Experimental evolution of microbial cells provides valuable information on evolutionary dynamics, such as mutations that contribute to fitness gain under given selection pressures. Although experimental evolution is a promising tool in evolutionary biology and bioengineering, long-term culture experiments under multiple environmental conditions often impose an excessive workload on researchers. Therefore, the development of automated systems significantly contributes to the advancement of experimental evolutionary research. This review presents several specialized devices designed for experimental evolution studies, such as an automated system for high-throughput culture experiments, a culture device that generate a temperature gradient, and an automated ultraviolet (UV) irradiation culture device. The ongoing development of such specialized devices is poised to continually expand new frontiers in experimental evolution research.

3D culture of ovarian follicles in granular and nanofibrillar hydrogels

3D culture of ovarian follicles in hydrogel matrices is an important emerging tool for basic scientific studies as well as clinical applications such as fertility preservation. For optimizing and scaling 3D culture of preantral follicles, there is a need for identifying biomaterial matrices that simplifies and improves the current culture procedures. At present, microencapsulation of follicles in alginate beads is the most commonly used approach. However, this technique involves notable manual handling and is best suited for encapsulation of single or several follicles. As a potential alternative, we here explore the suitability of different particle-based hydrogel matrices, where follicles can easily be introduced in tunable 3D environments, in large numbers. Specifically, we study the growth of secondary murine follicles in microgranular alginate and nanofibrillar cellulose matrices, with and without cell-binding cues, and map follicle growth against the viscoelastic properties of the matrices. We cultured follicles within the particle-based hydrogels for 10 days and continuously monitored their size, survival, and tendency to extrude oocytes. Interestingly, we observed that the diameter of the growing follicles increased significantly in the particle-based matrices, as compared to state-of-the-art alginate micro-encapsulation. On the other hand, the follicles displayed an increased tendency for early oocyte extrusion in the granular matrices, leading to a notable reduction in the number of intact follicles. We propose that this may be caused by impaired diffusion of nutrients and oxygen through thicker matrices, attributable to our experimental setup. Still, our findings suggest that viscoelastic, granular hydrogels represent promising matrices for 3D culture of early-stage ovarian follicles. In particular, these materials may easily be implemented in advanced culturing devices such as micro-perfusion systems.

Tape-assisted fabrication method for constructing PDMS membrane-containing culture devices with cyclic radial stretching stimulation.

Advanced in vitro culture systems have emerged as alternatives to animal testing and traditional cell culture methods in biomedical research. Polydimethylsiloxane (PDMS) is frequently used in creating sophisticated culture devices owing to its elastomeric properties, which allow mechanical stretching to simulate physiological movements in cell experiments. We introduce a straightforward method that uses three types of commercial tape-generic, magic and masking-to fabricate PDMS membranes with microscale thicknesses (47.2 µm for generic, 58.1 µm for magic and 89.37 µm for masking) in these devices. These membranes are shaped as the bases of culture wells and can perform cyclic radial movements controlled via a vacuum system. In experiments with A549 cells under three mechanical stimulation conditions, we analysed transcriptional regulators responsive to external mechanical stimuli. Results indicated increased nuclear yes-associated protein (YAP) and transcriptional coactivator with PDZ-binding motif (TAZ) activity in both confluent and densely packed cells under cyclically mechanical strains (Pearson's coefficient (PC) of 0.59 in confluent and 0.24 in dense cells) compared with static (PC = 0.47 in confluent and 0.13 in dense) and stretched conditions (PC = 0.55 in confluent and 0.20 in dense). This technique offers laboratories without microfabrication capabilities a viable option for exploring cellular behaviour under dynamic mechanical stimulation using PDMS membrane-equipped devices.

Development of a 3-dimensional organotypic model with characteristics of peripheral sensory nerves

We developed a rat dorsal root ganglion (DRG)-derived sensory nerve organotypic model by culturing DRG explants on an organoid culture device. With this method, a large number of organotypic cultures can be produced simultaneously with high reproducibility simply by seeding DRG explants derived from rat embryos. Unlike previous DRG explant models, this organotypic model consists of a ganglion and an axon bundle with myelinated A fibers, unmyelinated C fibers, and stereo-myelin-forming nodes of Ranvier. The model also exhibits Ca2+ signaling in cell bodies in response to application of chemical stimuli to nerve terminals. Further, axonal transection increases the activating transcription factor 3 mRNA level in ganglia. Axons and myelin are shown to regenerate 14days following transection. Our sensory organotypic model enables analysis of neuronal excitability in response to pain stimuli and tracking of morphological changes in the axon bundle over weeks.

Parylene C Coating Efficacy Studies: Enhancing Biocompatibility of 3D Printed Polyurethane Parts for Biopharmaceutical and CGT Applications.

Additive manufacturing, particularly Vat photopolymerization, presents a promising technique for producing complex, tailor-made structures, making it an attractive option for generating single-use components used in biopharmaceutical manufacturing equipment or cell culture devices. However, the potential leaching of cytotoxic compounds from Vat photopolymer resins poses a significant concern, especially regarding cell growth and viability in cell culture applications. This study explores the potential of parylene C coating to enhance the inertness of a polyurethane-based photopolymer resin, aiming to prevent cytotoxicity and improve biocompatibility. The study includes an analysis of extractables from the resin and photoinitiator to evaluate the resin's composition and to define selected marker compounds for investigating the coating efficiency. The time-dependent accumulation of relevant extractable compounds over a 70-day period are assessed to address the long-term use of the coated components. The impact of irradiation on the material and the coating was evaluated, along with an accelerated aging study to address the long-term performance of the coating. Biocompatibility in terms of in vitro cell growth studies is evaluated using Chinese hamster ovary cells, a standard cell line in biopharmaceutical manufacturing. Results demonstrate that parylene C coating significantly reduces the release of cytotoxic compounds, such as the photoinitiator diphenyl(2,4,6-trimethylbenzoyl)phosphine oxide (TPO). Although accelerated aging indicates a reduction in the barrier properties of the coating over time, the parylene C coating still effectively slows the release of extractables and significantly improves cell compatibility of the 3D printed parts. The findings suggest that parylene C-coated components can be safely integrated into biopharmaceutical manufacturing processes, with recommendations to minimize storage times between coating application and use to ensure optimal performance.

Rapid prototyping of perfusion cell culture devices for three-dimensional imaging of mesenchymal stem cell deposition and proliferation

Perfusion of porous scaffolds transports cells to the surface to yield cellular constructs for 3D models of disease and for tissue engineering applications. While ceramic scaffolds mimic the structure and composition of trabecular bone, their opacity and tortuous pores limit the penetration of light into the interior. Scaffolds that are both perfusable and amenable to fluorescence microscopy are therefore needed to visualize the spatiotemporal dynamics of cells in the bone microenvironment. In this study, a hybrid injection molding approach was designed to enable rapid prototyping of collector arrays with variable configurations that are amenable to longitudinal imaging of attached human mesenchymal stem cells (hMSCs) using fluorescence microscopy. Cylindrical collectors were arranged in an array that is permeable to perfusion in the xy-plane and to light in the z-direction for imaging from below. The effects of the collector radius, number, and spacing on the collection efficiency of perfused hMSCs was simulated using computational fluid dynamics (CFD) and measured experimentally using fluorescence microscopy. The effect of collector diameter on simulated and experimental cell collection efficiencies followed a trend similar to that predicted by interception theory corrected for intermolecular and hydrodynamic forces for the arrays with constant collector spacing. In contrast, arrays designed with constant collector number yielded collection efficiencies that poorly fit the trend with collector radius predicted by interception theory. CFD simulations of collection efficiency agreed with experimental measurements within a factor of two. These findings highlight the utility of CFD simulations and hybrid injection molding for rapid prototyping of collector arrays to optimize the longitudinal imaging of cells without the need for expensive and time-consuming tooling.

From Chips-in-Lab to Point-of-Care Live Cell Device: Development of a Microfluidic Device for On-Site Cell Culture and High-Throughput Drug Screening.

Live cell assays provide real-time data of cellular responses. In combination with microfluidics, applications such as automated and high-throughput drug screening on live cells can be accomplished in small devices. However, their application in point-of-care testing (POCT) is limited by the requirement for bulky equipment to maintain optimal cell culture conditions. In this study, we propose a POCT device that allows on-site cell culture and high-throughput drug screening on live cells. We first observe that cell viabilities are substantially affected by liquid evaporation within the microfluidic device, which is intrinsic to the polydimethylsiloxane (PDMS) material due to its hydrophobic nature and nanopatterned surface. The unwanted PDMS-liquid-air interface in the cell culture environment can be eliminated by maintaining a persistent humidity of 95-100% or submerging the whole microfluidic device under water. Our results demonstrate that in the POCT device equipped with a water tank, both primary cells and cell lines can be maintained for up to 1 week without the need for external cell culture equipment. Moreover, this device is powered by a standard alkali battery and can automatically screen over 5000 combinatorial drug conditions for regulating neural stem cell differentiation. By monitoring dynamic variations in fluorescent markers, we determine the optimal doses of platelet-derived growth factor and epidermal growth factor to suppress proinflammatory S100A9-induced neuronal toxicities. Overall, this study presents an opportunity to transform lab-on-a-chip technology from a laboratory-based approach to actual point-of-care devices capable of performing complex experimental procedures on-site and offers significant advancements in the fields of personalized medicine and rapid clinical diagnostics.

Evaluation of the effectiveness and user-friendliness of portable culture device for onsite detection of Salmonella spp., Listeria monocytogenes, Bacillus cereus and enumeration of total viable count from food samples

Evaluation of the effectiveness and user-friendliness of portable culture device for onsite detection of Salmonella spp., Listeria monocytogenes, Bacillus cereus and enumeration of total viable count from food samples

Remembered borders and senses of national belonging. Museums, monuments and commemorative sites as cultural artifacts and devices of power in Brazil, Paraguay and Argentina

Remembered borders and senses of national belonging. Museums, monuments and commemorative sites as cultural artifacts and devices of power in Brazil, Paraguay and Argentina

Advancing Tissue Culture with Light-Driven 3D-Printed Microfluidic Devices.

Three-dimensional (3D) printing presents a compelling alternative for fabricating microfluidic devices, circumventing certain limitations associated with traditional soft lithography methods. Microfluidics play a crucial role in the biomedical sciences, particularly in the creation of tissue spheroids and pharmaceutical research. Among the various 3D printing techniques, light-driven methods such as stereolithography (SLA), digital light processing (DLP), and photopolymer inkjet printing have gained prominence in microfluidics due to their rapid prototyping capabilities, high-resolution printing, and low processing temperatures. This review offers a comprehensive overview of light-driven 3D printing techniques used in the fabrication of advanced microfluidic devices. It explores biomedical applications for 3D-printed microfluidics and provides insights into their potential impact and functionality within the biomedical field. We further summarize three light-driven 3D printing strategies for producing biomedical microfluidic systems: direct construction of microfluidic devices for cell culture, PDMS-based microfluidic devices for tissue engineering, and a modular SLA-printed microfluidic chip to co-culture and monitor cells.