{"title":"Optical tweezer-assisted cell pairing and fusion for somatic cell nuclear transfer within an open microchannel.","authors":"Yidi Zhang, Han Zhao, Zhenlin Chen, Zhen Liu, Hanjin Huang, Yun Qu, Yaowei Liu, Mingzhu Sun, Dong Sun, Xin Zhao","doi":"10.1039/d4lc00561a","DOIUrl":"10.1039/d4lc00561a","url":null,"abstract":"<p><p>Somatic cell nuclear transfer (SCNT), referred to as somatic cell cloning, is a pivotal biotechnological technique utilized across various applications. Although robotic SCNT is currently available, the subsequent oocyte electrical activation/reconstructed embryo electrofusion is still manually completed by skilled operators, presenting challenges in efficient manipulation due to the uncontrollable positioning of the reconstructed embryo. This study introduces a robotic SCNT-electrofusion system to enable high-precision batch SCNT cloning. The proposed system integrates optical tweezers and microfluidic technologies. An optical tweezer is employed to facilitate somatic cells in precisely reaching the fusion site, and a specific polydimethylsiloxane (PDMS) chip is designed to assist in positioning and pairing oocytes and somatic cells. Enhancement in the electric field distribution between two parallel electrodes by PDMS pillars significantly reduces the required external voltage for electrofusion/electrical activation. We employed porcine oocytes and porcine fetal fibroblasts for SCNT experiments. The experimental results show that 90.56% of oocytes successfully paired with somatic cells to form reconstructed embryos, 76.43% of the reconstructed embryos successfully fused, and 70.55% of these embryos underwent cleavage. It demonstrates that the present system achieves the robotic implementation of oocyte electrical activation/reconstructed embryo electrofusion. By leveraging the advantages of batch operations using microfluidics, it proposes an innovative robotic cloning procedure that scales embryo cloning.</p>","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":" ","pages":"5215-5224"},"PeriodicalIF":6.1,"publicationDate":"2024-11-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142581171","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Lab on a ChipPub Date : 2024-11-07DOI: 10.1039/d4lc00659c
Claudia Allan, Yiling Sun, Stephen C. Whisson, Michael Porter, Petra C. Boevink, Volker Nock, Claudia-Nicole Meisrimler
{"title":"Observing root growth and signalling responses to stress gradients and pathogens using the bi-directional dual-flow RootChip","authors":"Claudia Allan, Yiling Sun, Stephen C. Whisson, Michael Porter, Petra C. Boevink, Volker Nock, Claudia-Nicole Meisrimler","doi":"10.1039/d4lc00659c","DOIUrl":"https://doi.org/10.1039/d4lc00659c","url":null,"abstract":"Plants respond to environmental stressors with adaptive changes in growth and development. Central to these responses is the role of calcium (Ca<small><sup>2+</sup></small>) as a key secondary messenger. Here, the bi-directional dual-flow RootChip (bi-dfRC) microfluidic platform was used to study defence signalling and root growth. By introducing salinity as sodium chloride (NaCl) treatment <em>via</em> a multiplexed media delivery system (MMDS), dynamic gradients were created, mimicking natural environmental fluctuations. Signal analysis in <em>Arabidopsis thaliana</em> plants showed that the Ca<small><sup>2+</sup></small> burst indicated by the G-CaMP3 was concentration dependent. A Ca<small><sup>2+</sup></small> burst initiated in response to salinity increase, specifically within the stele tissue, for 30 seconds. The signal then intensified in epidermal cells directly in contact with the stressor, spreading directionally towards the root tip, over 5 minutes. Inhibition of propidium iodide (PI) stain transport through the xylem was observed following salinity increase, contrasting with flow observed under control conditions. The interaction of <em>Phytophthora capsici</em> zoospores with <em>A. thaliana</em> roots was also studied. An immediate directional Ca<small><sup>2+</sup></small> signal was observed during early pathogen recognition, while a gradual, non-directional increase was observed in Orp1_roGFP fluorescent H<small><sub>2</sub></small>O<small><sub>2</sub></small> levels, over 30 min. By adjusting the dimensions of the bi-dfRC, plants with varying root architectures were subjected to growth analysis. Growth reduction was observed in <em>A. thaliana</em> and <em>Nicotiana benthamiana</em> roots when exposed to salinity induced by 100 mM NaCl, while <em>Solanum lycopersicum</em> exhibited growth increase over 90 minutes at the same NaCl concentration. Furthermore, novel insights into force sensing in roots were gained through the engineering of displaceable pillars into the bi-dfRC channel. These findings highlight the vital role of controlling fluid flow in microfluidic channels in advancing our understanding of root physiology under stress conditions.","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":"28 1","pages":""},"PeriodicalIF":6.1,"publicationDate":"2024-11-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142594925","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Microstring-engineered tension tissues: A novel platform for replicating tissue mechanics and advancing mechanobiology","authors":"Zixing Zhou, Tingting Li, Wei Cai, Xiaobin Zhu, Zuoqi Zhang, Guoyou Huang","doi":"10.1039/d4lc00753k","DOIUrl":"https://doi.org/10.1039/d4lc00753k","url":null,"abstract":"Replicating the mechanical tension of natural tissues is essential for maintaining organ function and stability, posing a central challenge in tissue engineering and regenerative medicine. Existing methods for constructing tension tissues often encounter limitations in flexibility, scalability, or cost-effectiveness. This study introduces a novel approach to fabricate soft microstring chips using a sacrificial template method, which is easy to operate, offers controlled preparation, and is cost-effective. Through experimental testing and finite element simulations, we validated and characterized the relationship between microstring deformation, tissue width, and the reaction force exerted by the microstrings, enabling precise measurement of tissue contraction force. We successfully constructed microstring-engineered tension tissues (METTs) and demonstrated that they exhibit a significant mechanical response to profibrotic factors. Additionally, we conceptually demonstrated the application of microstring chips in constructing METTs with asymmetric, biomimetic constraints. The results indicate effective construction and regulation of METTs, providing a robust platform for mechanobiology and biomedical research.","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":"1 1","pages":""},"PeriodicalIF":6.1,"publicationDate":"2024-11-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142588791","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Lab on a ChipPub Date : 2024-11-06DOI: 10.1039/d4lc00763h
Emma J.M. Moonen, Walther Verberne, Eduard Pelssers, Jason Heikenfeld, Jaap den Toonder
{"title":"Discretised microfluidics for noninvasive health monitoring using sweat sensing","authors":"Emma J.M. Moonen, Walther Verberne, Eduard Pelssers, Jason Heikenfeld, Jaap den Toonder","doi":"10.1039/d4lc00763h","DOIUrl":"https://doi.org/10.1039/d4lc00763h","url":null,"abstract":"Monitoring of chemical biomarker concentrations is often necessary in modern healthcare facilities, but it remains a challenge to do this frequently, minimally invasively to the patient, and fit to workflows of healthcare professionals. The use of sweat as a biofluid can address these issues. Unlike blood, sweat can be noninvasively and continuously sampled without direct involvement of a professional, and sweat contains a rich composition of biomarkers. However, patients in resting state have extremely low sweat rates and they produce correspondingly small sweat volumes, which makes sweat sensing of hospitalised patients highly challenging. We propose a unique solution that enables the use of sweat as a viable biofluid for noninvasive health monitoring, by actively transporting the sweat in a discretised manner. Our device uses electrowetting-on-dielectrics (EWOD) to create and move sweat droplets with a volume of around 1 nanolitre from a sweat gland to sensors integrated in the device. We present the first wearable device with integrated EWOD, and we show that it can collect and transport sweat on-body, while measuring sweat rate.","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":"95 1","pages":""},"PeriodicalIF":6.1,"publicationDate":"2024-11-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142594926","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"5-axis CNC micro-milling machine for three-dimensional microfluidics","authors":"Mitchell Modarelli, Devin Kot-Thompson, Kazunori Hoshino","doi":"10.1039/d4lc00496e","DOIUrl":"https://doi.org/10.1039/d4lc00496e","url":null,"abstract":"The gold standard of microfluidic fabrication techniques, SU-8 patterning, requires photolithography equipment and facilities and is not suitable for 3D microfluidics. A 3D printer is more convenient and may achieve high resolutions comparable to conventional photolithography, but only with select materials. Alternatively, 5-axis computer numerical control (CNC) micro-milling machines can efficiently prototype structures with high resolutions, high aspect ratios, and non-planar geometries from a variety of materials. These machines, however, have not been catered for laboratory-based, small-batch microfluidics development and are largely inaccessible to researchers. In this paper, we present a new 5-axis CNC micro-milling machine specifically designed for prototyping 3D microfluidic channels, made affordable for research and laboratories. The machine is assembled from commercially available products and custom-build parts, occupying 0.72 cubic meters, and operating entirely from computer aided design (CAD) and manufacturing (CAM) software. The 5-axis CNC micro-milling machine achieves sub-µm bidirectional repeatability (≤0.23 µm), machinable features <20 µm, and a work volume of 50 x 50 x 68 mm. The tool compatibility and milling parameters were designed to enable fabrication of virtually any mill-able material including metals like aluminum, brass, stainless steel, and titanium alloys. To demonstrate milling high resolution and high aspect ratios, we milled a thin wall from 360 brass with a width of 18.1 µm and an aspect ratio of ~50:1. We also demonstrated fabricating molds from 360 brass with non-planar geometries to create polydimethylsiloxane (PDMS) microfluidic channels. These included a channel on a 90° edge and a channel on a rounded edge with a 250-µm radius of curvature. Our 5-axis CNC micro-milling machine offers the most versatility in prototyping microfluidics by enabling high resolutions, geometric complexity, a large work volume, and broad material compatibility, all within a user-friendly benchtop system.","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":"214 1","pages":""},"PeriodicalIF":6.1,"publicationDate":"2024-11-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142574375","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Lab on a ChipPub Date : 2024-10-31DOI: 10.1039/d4lc00652f
Wali Inam, Anton Vladyka, joanna pylvanainen, junel solis, dado tokic, pasi kankaanpaa, Hongbo Zhang
{"title":"An imaging scheme to study the flow dynamics of Co-Flow regime in Microfluidics: Implications for Nanoprecipitation","authors":"Wali Inam, Anton Vladyka, joanna pylvanainen, junel solis, dado tokic, pasi kankaanpaa, Hongbo Zhang","doi":"10.1039/d4lc00652f","DOIUrl":"https://doi.org/10.1039/d4lc00652f","url":null,"abstract":"Co-flow microfluidics, in addition to its application in droplet generation, has gained popularity for use with miscible solvent systems (continuous microfluidics). By leveraging the small diffusional distances in miniature devices, processes like nanomaterial synthesis can be precisely tailored for high-throughput production. In this context, the manipulation of flow regimes—from laminar to vortex formation, as well as the generation of turbulent and turbulent jet flows—plays a significant role in optimizing these processes. Therefore, a detailed understanding of fluid interactions within microchannels is crucial. Imaging is a common approach to studying fluid behavior, often utilizing tracer particles. In search of alternative methodologies, we present a new imaging-based scheme to explore fluid interactions in various co-flow regimes through optical flow analysis, specifically using Gaussian window Mean Squared Error (MSE). By examining fluid flow characteristics such as flow intensities (caused by fluctuations) and the projected movement of fluid spots, we characterize slow vortexing and chaotic flow behaviors in co-flow regimes. Consequently, we use imaging data to illustrate the influence of co-flow regimes on particle synthesis. This new tool provides the scientific community with an innovative method to study fluid interactions, which can be further explored to develop a more effective understanding of fluid mixing and optimize fluid manipulation in microfluidic devices","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":"4 1","pages":""},"PeriodicalIF":6.1,"publicationDate":"2024-10-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142556238","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Lab on a ChipPub Date : 2024-10-30DOI: 10.1039/d4lc00721b
Zhifang Liu, Yuwen Lu, Wei Tan, Guorui Zhu
{"title":"Dual-drive acoustic micromixer for rapid nucleation and ultrafast growth of perovskite nanoparticles","authors":"Zhifang Liu, Yuwen Lu, Wei Tan, Guorui Zhu","doi":"10.1039/d4lc00721b","DOIUrl":"https://doi.org/10.1039/d4lc00721b","url":null,"abstract":"All-inorganic cesium lead halide perovskites have garnered significant attention owing to their favorable optical properties. Microfluidics-based acoustic mixers are capable of achieving rapid nucleation and ultrafast growth kinetics. Nevertheless, conventional acoustic mixers rely on the response of microstructures to the acoustic field for mixing fluids, the majority of these disturbances occur in the central region of the channel, with minimal impact on the fluid within the side walls. This paper proposes a novel acoustic mixer that combines the effects of sharp corners and bubbles in response to the acoustic field, thereby producing effective disturbance of the fluid throughout the channel. The combined effect enables the micromixer to achieve complete 2 mixing at different inlet flow ratios with mixing times as low as 5 ms. The superiority of acoustic mixers in controlling the nanocrystal formation stage was further validated through the synthesis of chalcogenide nanocrystals using the LARP method. The millisecond mixing time facilitated the rapid formation of nanocrystals and their subsequent rapid growth. The results demonstrate that the green luminescence intensity at 520 nm of the samples synthesized by the acoustic micromixer is 118% higher than that of the samples synthesized by the intermittent reactor. The novel micromixer broadens the range of applications and offers a promising avenue for the large-scale continuous synthesis of high-quality lead-halide perovskite nanocrystals (NCs).","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":"8 1","pages":""},"PeriodicalIF":6.1,"publicationDate":"2024-10-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142556239","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Lab on a ChipPub Date : 2024-10-25DOI: 10.1039/d4lc00205a
Chase Paterson, Elizabeth Vargis
{"title":"Applying Low Levels of Strain to Model Nascent Phenomenon of Retinal Pathologies","authors":"Chase Paterson, Elizabeth Vargis","doi":"10.1039/d4lc00205a","DOIUrl":"https://doi.org/10.1039/d4lc00205a","url":null,"abstract":"Age-related macular degeneration (AMD) is a leading cause of vision loss in the elderly. A better understanding of the mechanisms of the disease, especially at early stages, could elucidate new treatment targets. One characteristic of AMD is strain on the retinal pigment epithelium (RPE), a crucial layer of the retina. This strain can be caused by physical phenomena like waste aggregation underneath the RPE from aging, drusen formation, or leaky blood vessels that infiltrate the retina during choroidal neovascularization (CNV). It is not well understood how strain affects RPE cells. Most models generate equibiaxial strain or higher levels of strain that are not representative of early stages of AMD. To overcome these issues, we have engineered a device to cause controlled, low amounts of localized, radial strain (maximum ~2%). This strain level is more mimetic to what occurs during aging or at the beginning of physical disruptions experienced during AMD. To evaluate how RPE cells respond to this physical stimulus, primary porcine RPE cells were exposed to low levels of strain applied by our custom-made device. Cell secretions and genetic expression were analyzed to see how proteins linked to drusen and CNV are affected. The results indicate that this low amount of strain does not immediately initiate angiogenesis but causes changes in mRNA expression of amyloid precursor protein (APP), which plays a role in retinal health and drusen accumulation. This research offers insight into AMD progression as well as the health of other organs, including the brain.","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":"212 1","pages":""},"PeriodicalIF":6.1,"publicationDate":"2024-10-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142490256","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"A Gravity-Driven Tissue Chip to Study Efficacy and Toxicity of Cancer Therapeutics","authors":"Pouria Rafsanjani Nejad, Astha Lamichhane, Prasiddha Guragain, Gary Luker, Hossein Tavana","doi":"10.1039/d4lc00404c","DOIUrl":"https://doi.org/10.1039/d4lc00404c","url":null,"abstract":"Tissue chip technologies have emerged as promising tools in preclinical studies. In oncology, this is driven by the high failure rates of candidate drugs in clinical trials mainly due to inadequate efficacy or intolerable toxicity and the need for better predictive preclinical models than those traditionally used. However, the intricate design, fabrication, operation, and limited compatibility with automation limit the utility of tissue chips. To tackle these issues, we designed a novel 32-unit tissue chip in the format of standard 96-well plates to streamline automation, fabricated it using 3D printing, and leveraged gravity-driven flow to bypass the need for external flow devices. Each unit includes three interconnected tissue compartments that model liver, tumor, and bone marrow stroma. Focus on liver and bone marrow stroma was due to their respective roles in drug metabolism and disturbances to the bone marrow niche from off-target toxicity of chemotherapies. We analyzed flow patterns, mixing, and oxygen transport among and within the compartments through finite element simulations and demonstrated the utility of the tissue chip to study the efficacy of commonly-used cytotoxic cancer drugs against tumor cells and their toxicity toward liver and bone marrow cells.","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":"10 1","pages":""},"PeriodicalIF":6.1,"publicationDate":"2024-10-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142489901","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}