HardwareX

A very large number of biology and biochemistry laboratory protocols require transferring liquid aliquots from individual containers into individual wells of a multi-well plate, from plates to individual containers, or from one plate to another. Doing this by hand without errors, such as skipping wells, placing two samples in the same well, or swapping sample locations, especially when using plates with 96 wells or more, is difficult and requires enormous operator focus and/or a tedious manual error checking system. We present here a device built to facilitate error-free pipetting of samples from individual barcoded tubes to a multi-well plate or between multi-well plates (both 96 and 384 wells are supported). The device is programmable, modular and easily customizable to accommodate plates with different form-factors, and different protocols. The main components are only a 12.3" touch screen, a small form-factor PC, and a barcode scanner, combined with custom-made parts can be easily fabricated with a laser cutter and a hobby-grade 3D printer. The total cost is between approximately US$550 and US$600, depending on the configuration. Specifications table O_TBL View this table: org.highwire.dtl.DTLVardef@8554f4org.highwire.dtl.DTLVardef@18c8878org.highwire.dtl.DTLVardef@153b050org.highwire.dtl.DTLVardef@15c7de6org.highwire.dtl.DTLVardef@14daeb7_HPS_FORMAT_FIGEXP M_TBL C_TBL

Measuring optical density (OD) is a very common technique in biological laboratories to determine the concentration of a substance in solution or of bacteria (or microscopic particles) in suspension. For example, bacterial cultures engineered to produce (express) a protein or compound of interest are a workhorse of modern molecular biology laboratories. Commonly, the expression of the product is triggered (induced) by a chemical signal added to the culture at the proper time in the growth curve of the culture (typically in the middle of the exponential growth phase, at an OD value of [~]0.6). The most common tool for measuring OD is a spectrophotometer. However, most spectrophotometers are sophisticated, non-portable and expensive laboratory instruments, costing tens of thousands of dollars. Even a very low cost spectrophotometer for educational use costs at least US$1,000. Because of the cost, even well resourced labs have only one instrument, which becomes a bottleneck when multiple bacterial cultures need to be monitored simultaneously. The problem is more acute in developing countries, where multiple labs have to share a single spectrophotometer, or theres no such instrument at all. Having a cheap and simple device to measure OD would enable multiple people in a laboratory to monitor their bacterial cultures independently, even in resource-limited settings. At the same time, a portable OD meter could be useful for field work. Here we present the detailed build instructions and characterization of a very simple OD meter that costs only US$60, and can measure OD values from [~]0.05 to 2.0. Specifications table O_TBL View this table: org.highwire.dtl.DTLVardef@721005org.highwire.dtl.DTLVardef@79c5d3org.highwire.dtl.DTLVardef@aabb95org.highwire.dtl.DTLVardef@1016f09org.highwire.dtl.DTLVardef@120f3e2_HPS_FORMAT_FIGEXP M_TBL C_TBL

We report an automated, low-cost, and open-source methodology for rodent object recognition tasks (ORT), which are widely used behavior paradigms. The system was designed with an effort of behavior standardization and improvement. When it combined with easily accessible 3-D manufacturing technology, this proximity capacitive sensor based method provides an opportunity to explore new, and high throughput hardware for NORT. The systems software allows behavior data to be instantaneously monitored and its output can be plotted promptly immediately upon the completion of the behavior test, thus saving labor and time. The performance of this package is cross-verified with a widely used video-based automated software scoring method and examined its sensitivity to chemogenetic behavior manipulations. This integrated package is an affordable and accurate tool for translational recognition memory research and other type of research with frequent use of NORT paradigms. Visual Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=52 SRC="FIGDIR/small/518044v1_ufig1.gif" ALT="Figure 1"> View larger version (18K): org.highwire.dtl.DTLVardef@1205ca6org.highwire.dtl.DTLVardef@1a2d156org.highwire.dtl.DTLVardef@5d8ef9org.highwire.dtl.DTLVardef@5589ee_HPS_FORMAT_FIGEXP M_FIG C_FIG

Microfluidic systems are widely used in biology for their ability to control environmental parameters. Specifically, cell culture or chemistry in microfluidic devices requires tight control of the temperature. In addition, microfluidic devices can be made transparent to visible light and compatible with inverted microscopes. Yet, the current temperature control systems that allow high-resolution microscopy either require a set of complex secondary channels, a bulky, expensive, and microscope-dependent incubator, or fail to produce a homogenous temperature profile across the sample area. Here, we present HeatChips, a simple, cost-effective system to heat samples inside PDMS-based microfluidic devices in a homogeneous manner. It is based on a transparent heating glass in contact with the top of the microfluidic device, and a contactless, infrared temperature sensor attached to the objective that directly reads the temperature of the bottom of the chip. This portable system is compatible with most chip designs and allows imaging of the sample on inverted microscopes for extended periods of time without any optical restriction, for a cost of less than 100{euro}. Specifications table [Table 1]

Drug testing is a vital step in identification of the potential efficacy of any new/existing drug and/or combinations of drugs. The conventional methods of testing the efficacy of new drugs using multi-well plates are time consuming, prone to evaporation loss and manual error. Microfluidic devices with automated generation of concentration gradient provide a promising alternative. The implementation of such microfluidic devices is still limited owing to the additional expertise and facilities required to fabricate and run these devices. Conventional microfluidic devices also need pumps, tubings, valves, and other accessories, making them bulky and nonportable. To address these problems, we have developed a method for fabricating microfluidic structures using a nonconventional technique by exploiting the Saffman-Taylor instability in lifted Hele-Shaw cell. Multi-channel structure molds with varying dimensions were fabricated by shaping ceramic polymer slurry and retaining the shape. Further using the mold thus made, polydimethyl siloxane (PDMS) devices offering static, stable, diffusion-based gradient were casted using soft lithography. We have demonstrated with COMSOL simulation, as well as using Fluorescein isothiocyanate (FITC), a fluorescent dye, that the concentration gradient can be generated in this device, which remains stable for at least 5 days. Using this multichannel device, in vitro drug efficacy was validated with two drugs namely-Temozolomide (TMZ) and Curcumin, one FDA approved and one under research, on glioblastoma cells (U87MG). The resulting IC50 values were consistent with those reported in literature. We have also demonstrated the possibility of conducting molecular assays post-drug testing in the device by microtubule staining after curcumin treatment on cervical cancer cells (HeLa). In summary, we have demonstrated a i) user-friendly, ii) portable, static drug testing platform that iii) does not require further accessories and can create iv) a stable gradient for long duration. Such a device can reduce the time, manual errors, fabrication and running expenditure, and resources to a great extent in drug testing. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=142 SRC="FIGDIR/small/505813v1_ufig1.gif" ALT="Figure 1"> View larger version (79K): org.highwire.dtl.DTLVardef@e1f648org.highwire.dtl.DTLVardef@f029d7org.highwire.dtl.DTLVardef@14afd68org.highwire.dtl.DTLVardef@42b888_HPS_FORMAT_FIGEXP M_FIG C_FIG

Planktonic organisms are a cornerstone of marine ecosystems. They vary significantly in size and have a repertoire of behaviors to aid them to survive and navigate their three-dimensional environment. One of the most important cues is light. A variety of setups were used to study the swimming behavior of specific organisms, but broader and comparative investigations need more versatile solutions. With the help of 3D printing, we designed and constructed a modular and flexible behavioral observation setup that enables recordings of animals down to 50m or up to a few centimeters. A video analysis pipeline using ImageJ and python allows a quick, automated, and robust tracking solution, capable of processing many videos automatically. A modular light path allows the addition of filters or use of pulse width modulation to equalize photon emission of LEDs or additional LEDs to mix different wavelengths. Optionally, a spectrometer can be installed to enable live monitoring of a stimulus. We tested the setup with two phototactic marine planktonic larvae. First, we investigated the spectral sensitivity of the 7-day old larvae of the polychaete Malacoceros fuliginosus and second, the behavior of the 200m spherical bryozoan coronated larvae of Tricellaria inopinata to ultraviolet light coming from the bottom of the vessel. The setup and pipeline were able to record and analyze hundreds of animals simultaneously. We present an inexpensive, modular, and flexible setup to study planktonic behavior of a variety of sizes.

We used DNA origami to create NanoNERF, the worlds smallest NERF blaster replica (Figure 1). We based our design on the NERF model Maverick Rev-6, and scaled the dimensions down three million times. NanoNERF is planar and measures [~]100 nm in length, with a length-to-width ratio closely resembling the original toy. Here, we describe the design, prototyping, and validation pipeline used to create the NanoNERF. We also discuss potential applications to motivate the creation of future nanoscale blasters with a firing functionality. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=41 SRC="FIGDIR/small/560388v3_fig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@1d7619forg.highwire.dtl.DTLVardef@14a16dforg.highwire.dtl.DTLVardef@123b531org.highwire.dtl.DTLVardef@14cb703_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 1:C_FLOATNO A nanoscale replica of the NERF Maverick Rev-6 a) Original NERF Maverick Rev-6 toy, b) rendering of the NanoNERF structure in oxView. Gray lines represent individual DNA strands. NanoNERF length: 100 nm; width of barrel: 35 nm; thickness: 2 nm; c) Scan of a NanoNERF blaster acquired in an Atomic Force Microscope. C_FIG

3D printed laboratory tools are a fast-developing field, and microscopes in particular have seen several high-quality projects reported in recent years. However, currently available projects for 3D printed microscopes have not been optimised for molecular biology applications. We present an open-source, 3D-printable optical microscope that has been specifically designed for affordability, ease of use, and versatility in molecular biology applications, particularly for fluorescence detection, time lapse imaging, and higher throughput scanning of microtitre plates. The microscope integrates a large computer numerically controlled (CNC) stage, allowing precise scanning of wells in standard microtiter plates, and is equipped with a Raspberry Pi HQ camera module and customizable optical components, including standard microscope objectives. The modular design allows for a range of illumination setups, such as a 3 W blue LED for basic epifluorescence, and supports additional modifications, including dark field or RGB matrix lighting. The design opens new possibilities for cheap home-built microscopes for bioscience applications that are modular and easily adaptable for specific user needs.

Live imaging of human or other mammalian cells at multi-hour time scales with minimal perturbation to their growth state requires that the specimens optimal growth conditions are met while fixed to a microscope stage. In general, ideal conditions include culturing in complete growth media, an ambient temperature of 36-37 C, and a humidity-controlled atmosphere comprising typically 5-7% CO2. Commercially available devices that achieve these conditions are not a financially viable option for many labs, with the price ranging anywhere from $12000 to $40000. The advent of 3D printing technology has allowed for low-cost rapid prototyping with precision comparable to traditional fabrication methods, opening the possibility for in-lab design and production of otherwise prohibitively expensive equipment such as stage-top incubation devices. The continued usefulness and widespread availability of single-board computers (SBC) such as Arduino and Raspberry Pi also simplify the process by which these devices can be controlled. Here we report the production of a do-it-yourself (DIY) device for stage-top incubation with temperature and atmospheric control with a cost reduction of approximately 100x.

The expanding scope of laboratory automation increasingly demands systems that can be tailored to specific experimental constraints, including footprint, timing, cost, and control. While open-source software has improved protocol flexibility, liquid-handling hardware itself remains largely closed, limiting the ability of academic and startup laboratories to build instruments around biological requirements rather than vendor defaults. Here, we present a fully open-source, purpose-built liquid-handling robot assembled from commercially available components and developed entirely in a research setting. The platform integrates open hardware, electronics, and a Python-based control stack compatible with PyLabRobot, exposing low-level motion dynamics and liquid-handling behaviors directly to experiment code. We validate the system using a high-throughput turbidostat workflow that requires rapid, closed-loop measurement and actuation to maintain microbial cultures at defined density setpoints. The robot sustains stable steady-state growth across approximately 200 cultures with heterogeneous growth dynamics. A replica build completed by two lab members in approximately one week confirms that the platform can be reproduced from its bill of materials and assembly guide. Its compact footprint and use of off-the-shelf components make it suitable for rapid, parallel deployment in settings such as public health emergencies or by distributed laboratories. Together, these results demonstrate that industry-class liquid handlers can be custom-built for specific experimental goals, establishing a blueprint for open, purpose-driven hardware development across research and industrial automation contexts. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=132 SRC="FIGDIR/small/709168v1_ufig1.gif" ALT="Figure 1"> View larger version (63K): org.highwire.dtl.DTLVardef@1b2cb4eorg.highwire.dtl.DTLVardef@1418e8dorg.highwire.dtl.DTLVardef@f60618org.highwire.dtl.DTLVardef@a3d3b_HPS_FORMAT_FIGEXP M_FIG Open Liquid Handler (OLH) Design Goals. Left: Design goals for a purpose-built platform for time-sensitive, closed-loop biological workflows, emphasizing high-accuracy dosing (low variability liquid handling), rapid integrated measurement (plate deck and isolated workspace), customizable deck and peripheral options, compact footprint with high throughput, containment via an enclosed wet workspace for biosafety and sterility, and a replicable build using off-the-shelf OEM components with open design files. Right: Open Liquid Handler design and physical implementation, with aerial and front views highlighting the enclosed cabinet and the working envelope over a compact deck. C_FIG

1Isothermal amplification-based methods for pathogen DNA or RNA detection offer high sensitivity, rapid detection, and the potential for deployment in remote fields and home testing. Consequently, they are emerging as alternatives to PCR and saw a surge in research activity and deployment for the rapid detection of SARS-CoV-2 during the Covid-19 pandemic. The most common isothermal DNA detection methods rely on minimal reagents for DNA amplification and simple hardware that can maintain isothermal conditions and read-out a fluorescent or colorimetric signal. Many researchers globally are working on improving these components based on diverse end-user needs. In this work, we have recognized the need for an open-source hardware device for isothermal amplification, composed of off-the-shelf components that are easily accessible in any part of the world, is easily manufacturable in a distributed and scalable way using 3D printing, and that can be powered using a wide diversity of batteries and power sources. We demonstrate the easy assembly of our device design and demonstrate its efficacy using colorimetric LAMP for both RNA and DNA targets.

Live-cell microscopy imaging typically involves the use of high-quality glass-bottom chambers that allow cell culture, gaseous buffer exchange and optical properties suitable for microscopy applications. However, commercial sources of these chambers can add significant annual costs to cell biology laboratories. Consumer products in three-dimensional printing technology, for both Filament Deposition Modeling (FDM) and Masked Stereo Lithography (MSLA), have resulted in more biomedical research labs adopting the use of these devices for prototyping and manufacturing of lab plastic-based items, but rarely consumables. Here we describe a modular, live-cell chamber with multiple design options that can be mixed per experiment. Single reusable carriers and the use of biodegradable plastics, in a hybrid of FDM and MSLA manufacturing methods, reduce plastic waste. The system is easy to adapt to bespoke designs, with concept-to-prototype in a single day, offers significant cost savings to the users over commercial sources, and no loss in dimensional quality or reliability.

Stable and productive CHO cell lines are essential for biopharmaceutical manufacturing, yet early expansion steps are often constrained by prolonged period required for suspension adaptation. Single-cell cloning (SCC) ensures monoclonality and regulatory compliance, but cells transitioning from static to suspension culture frequently exhibit variable recovery, which prolongs timelines and increases process variability. To address this challenge, mixing-based microplate culture systems have been developed to improve early expansion efficiency. The C.NEST platform provides controlled pneumatic mixing and environmental monitoring that facilitates earlier adaptation to suspension conditions. At the 96-well and 24-well stages, this system allows cells to establish stable growth under suspension-like environments, thereby shortening the adaptation period following transfer to shaking culture. In this study, we applied C.NEST to the SCC workflow for developing CHO-K1 stable cell lines. Integrating C.NESTs controlled mixing reduced adaptation time, enhanced the consistency of clone expansion, and improved the ability to identify high-yield clones. These findings highlight the potential of C.NEST to streamline cell line development workflows by accelerating early suspension adaptation and improving clone selection reliability. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=165 SRC="FIGDIR/small/693844v1_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@ca76bforg.highwire.dtl.DTLVardef@3a3079org.highwire.dtl.DTLVardef@447e01org.highwire.dtl.DTLVardef@ac8d6f_HPS_FORMAT_FIGEXP M_FIG C_FIG C.NEST mixing shortens suspension adaptation, accelerates clone expansion, and enhances early-stage screening. HighlightO_LIThe C.NEST microplate agitation culture system accelerates early CHO-K1 cell line development. C_LIO_LIControlled pneumatic mixing improved oxygen transfer and medium homogeneity, promoting stable growth during early expansion. C_LIO_LIEarly mixing shortens suspension adaptation by approximately one week. C_LIO_LIMixing cultures enabled more accurate clone performance assessment, revealing high-producing outliers. C_LIO_LIC.NEST provides a scalable and reproducible solution for integrating mixing-based culture into single-cell cloning workflows. C_LI

We present the BOOPTHAT (Batch-Operating Optically Powered Targeted Heater for Activating Transgenes) as a low-cost system for activating heat-shock inducible transgenes with spatiotemporal control in multiple zebrafish embryos at a time. Gaining finer spatiotemporal control over gene expression is critical for unraveling the regulatory networks that coordinate embryogenesis. While the heat-shock inducible gene expression system is a widely used tool for controlling temporal transgene expression, its applicability in spatiotemporal control is limited. By adding a level of spatial control onto this well-established system, we take advantage of the existing infrastructure surrounding the HS induction system and introduce new ways to use the multitude of existing lines. The BOOPTHAT system is built from 3D printed components and inexpensive consumer parts. Independent 3D printed micromanipulators are used to position optical fiber probes. When coupled to a light source, the probes are heated photothermally and are used to perform targeted gene activation in multiple samples at a time. We demonstrate the capabilities of our system and highlight some areas of research that stand to benefit from this frugal and effective system.

A variety of costly research-grade imaging devices are available for the detection of spectroscopic features. Here we present an affordable, open-source and versatile device, suitable for a range of applications. We provide the files to print the imaging chamber with commonly available 3D printers and instructions to assemble it with easily available hardware. The imager is suitable for rapid sample screening in research, as well as for educational purposes. We provide details and results for an already proven set-up which suits the needs of a research group and students interested in UV-induced near-infrared fluorescence detection of microbial colonies grown on Petri dishes. The fluorescence signal confirms the presence of bacteriochlorophyll a in aerobic anoxygenic phototrophic bacteria (AAPB). The imager allows for the rapid detection and subsequent isolation of AAPB colonies on Petri dishes with diverse environmental samples. To this date, 15 devices have been build and more than 7000 Petri dishes have been analyzed for AAPB, leading to over 1000 new AAPB isolates. Parts can be modified depending on needs and budget. The latest version with automated switches and double band pass filters costs around 350{euro} in materials and resolves bacterial colonies with diameters of 0.5 mm and larger. The low cost and modular build allow for the integration in high school classes to educate students on light properties, fluorescence and microbiology. Computer-aided design of 3D-printed parts and programming of the employed Raspberry Pi computer could be incorporated in computer sciences classes. Students have been also inspired to do agar art with microbes. The device is currently used in seven different high schools in Finland. Additionally, a science education network of Finnish universities has incorporated it in its program for high school students. Video guides have been produced to facilitate easy operation and accessibility of the device. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=62 SRC="FIGDIR/small/543100v2_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@6dfcb7org.highwire.dtl.DTLVardef@ea7fbcorg.highwire.dtl.DTLVardef@1681e78org.highwire.dtl.DTLVardef@a88d51_HPS_FORMAT_FIGEXP M_FIG C_FIG

Time-lapse imaging of bacteria growing in micro-channels in a controlled environment has been instrumental in studying the single cell dynamics of bacterial growth. This kind of a microfluidic setup with growth chambers is popularly known as mother machine [1]. In a typical experiment with such a set-up, bacterial growth can be studied for numerous generations with high resolution and temporal precision using image processing. However, as in any other experiment involving imaging, the image data from a typical mother machine experiment has considerable intensity fluctuations, cell intrusion, cell overlapping, filamentation etc. The large amount of data produced in such experiments makes it hard for manual analysis and correction of such unwanted aberrations. We have developed a modular code for segmentation and analysis of mother machine data (SAM) for rod shaped bacteria where we can detect such aberrations and correctly treat them without manual supervision. We track cumulative cell size and use an adaptive segmentation method to avoid faulty detection of cell division. SAM is currently written and compiled using MATLAB. It is fast ([~] 15 min/GB of image) and can be efficiently coupled with shell scripting to process large amount of data with systematic creation of output file structures and graphical results. It has been tested for many different experimental data and is publicly available in Github.

Developing a novel microfluidic organoid system required many experiments and iterations due to lack of knowledge about the relevant developmental biology. Collecting data on the developing organoids quickly escalated into a bottleneck as high throughput and long term culture resulted in a rapidly increasing number of specimens being observed. Commercially available automated microscope systems exist, but were either too expensive or not appropriate, and could not be modified. To satisfy the increasing need for automated data collection, a custom robotic system was developed to collect data from within a standard incubator. An X-Y belt driven gantry was designed with an architecture chosen to balance high accuracy, low cost, speed, and range of motion. Focus control was implemented with dual miniature leadscrews. A linear sliding mechanism was used to switch between two microscope objectives. 3D printed chip attachments were designed to implement illuminators for bright field imaging, and electrodes for stimulating the cardiac organoids. A fluorescent filter block was designed using a 3D printed piece to hold optical components, and a multi-band filter set that allowed for three color fluorescence without moving parts. A pulley driven tilting stage gravitationally biased the organoids during development. In order to ensure accurate image collection despite the inevitable position shifting of the chips, an image processing pipeline was developed for locating organoids using geometrical microfluidic chip features. The resulting robotic system automated imaging data collection on organoids and electrical and mechanical stimulation, in addition to being modifiable for future projects.

Collecting cells from zebrafish embryos for genotyping is critical to rapid research with these model organisms. The standard collection process is manual, labor-intensive, time-consuming, and requires a skilled person to perform it. To overcome this challenge, researchers are exploring the development of automated genotyping tools for live animals, which would significantly enhance the efficiency and accuracy of genetic screening in zebrafish and other species. The focus of this research was to optimize the Zebrafish Embryo Genotyper (ZEG), an automated system used for the rapid extraction of cellular material from zebrafish embryos. This system rapidly vibrates a roughened chip containing a zebrafish embryo to collect genetic material safely and efficiently. The aim was to improve the efficiency of DNA collection from the chips used with the ZEG by identifying the key factors that contribute to the process. First, the chips were modified to resolve issues associated with loss of sample volume from the chip wells due to evaporation during processing. Second, we experimented with three critical parameters - sample volume in the wells, the vibrational frequency of the system, and the operation time - on the quantity of DNA collected. The performance was evaluated by measuring embryo survival and quantifying the DNA collected. The sensitivity (previously 90%) of the DNA collection and embryo survival (previously 95%) of the were both found to be greater than 95% after optimization. The optimized design parameters (15 {micro}L solution volume, 2.4 V, and a 5-minute run with 5 s alternating on/off) provided a >50% increase in DNA collection compared to the previous designs and parameters. The proposed chip design and operation do not appear to cause any apparent adverse effects on the development or survival of the embryos.

BackgroundO_LIMany Drosophila species use acoustic communication during courtship and studies of these communication systems have provided insight into neurobiology, behavioral ecology, ethology, and evolution. C_LIO_LIRecording Drosophila courtship sounds and associated behavior is challenging, especially at high throughput, and previously designed devices are relatively expensive and complex to assemble. C_LI ResultsO_LIWe present construction plans for a modular system utilizing mostly off-the-shelf, relatively inexpensive components that provides simultaneous high-resolution audio and video recording of 96 isolated or paired Drosophila individuals. C_LIO_LIWe provide open-source control software to record audio and video. C_LIO_LIWe designed high intensity LED arrays that can be used to perform optogenetic activation and inactivation of labelled neurons. C_LIO_LIThe basic design can be modified to facilitate novel study designs or to record insects larger than Drosophila. C_LIO_LIFewer than 96 microphones can be used in the system if the full array is not required or to reduce costs. C_LI ImplicationsO_LIOur hardware design and software provide an improved platform for reliable and comparatively inexpensive high-throughput recording of Drosophila courtship acoustic and visual behavior and perhaps for recording acoustic signals of other small animals. C_LI

New in vitro models are an urgent need for to improve both the research data and the preclinical development of new drugs. Current standardized cellular models are mainly based in 2D cell culture, which lacks flow conditions and with complex co-culture settings. In this way, advanced cell culture models, such as organ-on-a-chip (OoC), aim to solve these limitations. OoC systems are composed by a microfluidic chip functionalized with different combinations of extracellular matrixes, coatings and cell cultures to mimic the physiological conditions of human organs. Advantages of OoC include the possibility to add 3D structures, delimited regions for co-culture and dynamic flow conditions to cell cultures. However, to perform reproducible and controlled experiments with OoC, it is necessary to systematically standardize the cell culture conditions in the microfluidic channels. For this is necessary to test both the combination of flow and extracellular matrix (ECM) coating to reliably mimic the human organ physiology. In this work, we standardized both conditions, ECM coating and the flow conditions to functionalize OoC with cell lines from kidney (Caki-1) and from lung (A549) to develop OoC systems beyond the Vessel-on-a-chip setting. In this way, the protocol detailed in this work will allow to standardize cell culture on different optimized OoC types with different cell types from different origins.