Alexis Lefevre, Cristian Brandi, Adele De Ninno, Filippo Ruggiero, Enrico Verona, Michael Gauthier, Paolo Bisegna, Aude Bolopion, Federica Caselli
This work presents an innovative all-electrical platform for selective single-particle manipulation. The platform combines microfluidic impedance cytometry for label-free particle characterization and dielectrophoresis for contactless multi-way particle separation. The microfluidic chip has a straightforward coplanar electrode layout and no particle pre-focusing mechanism is required. An original online algorithm analyzes the impedance signals of each incoming particle and regulates in real-time the dielectrophoretic voltages according to a desired control logic. As proof-of-concept, three operation modes are demonstrated on a mixture of 8, 10, and 12 µm diameter beads: (i) particle position swapping across channel axis, irrespective of particle size, (ii) size-based particle separation, irrespective of particle position, and (iii) sorting of a selected sequence of particles. As a perspective, the versatility of impedance cytometry and dielectrophoresis and the possibility to configure alternative control logics hold promises for advanced particle and cell manipulation.
{"title":"Real-time impedance-activated dielectrophoretic actuation for reconfigurable manipulation of single flowing particles","authors":"Alexis Lefevre, Cristian Brandi, Adele De Ninno, Filippo Ruggiero, Enrico Verona, Michael Gauthier, Paolo Bisegna, Aude Bolopion, Federica Caselli","doi":"10.1039/d4lc00622d","DOIUrl":"https://doi.org/10.1039/d4lc00622d","url":null,"abstract":"This work presents an innovative all-electrical platform for selective single-particle manipulation. The platform combines microfluidic impedance cytometry for label-free particle characterization and dielectrophoresis for contactless multi-way particle separation. The microfluidic chip has a straightforward coplanar electrode layout and no particle pre-focusing mechanism is required. An original online algorithm analyzes the impedance signals of each incoming particle and regulates in real-time the dielectrophoretic voltages according to a desired control logic. As proof-of-concept, three operation modes are demonstrated on a mixture of 8, 10, and 12 µm diameter beads: (i) particle position swapping across channel axis, irrespective of particle size, (ii) size-based particle separation, irrespective of particle position, and (iii) sorting of a selected sequence of particles. As a perspective, the versatility of impedance cytometry and dielectrophoresis and the possibility to configure alternative control logics hold promises for advanced particle and cell manipulation.","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":null,"pages":null},"PeriodicalIF":6.1,"publicationDate":"2024-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142431705","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}
With advancements in human induced pluripotent stem cell (hiPSC) technology, there is an increasing demand for quality control techniques to manage the long-term process of target cell production effectively. While monitoring systems designed for use within incubators are promising for assessing culture quality, existing systems still face challenges in terms of compactness, throughput, and available metrics. To address these limitations, we have developed a compact and high-throughput lens-free imaging device named INSPCTOR. The device is as small as a standard culture plate, which allows for the installation of multiple units within an incubator. INSPCTOR utilises a large thin-film transistor image sensor, enabling simultaneous observation of six independent culture environments, each approximately 1 cm2. With this device, we successfully monitored the confluency of hiPSC cultures and identified the onset timing of epithelial-to-mesenchymal transition during mesodermal induction. Additionally, we quantified the beating frequency and conduction of hiPSC-derived cardiomyocytes by using high-speed imaging modes. This enabled us to identify the onset of spontaneous beating during differentiation and assess chronotropic responses in drug evaluations. Moreover, by tracking beating frequency over 10 days of cardiomyocyte maturation, we identified week-scale and daily-scale fluctuations, the latter of which correlated with cellular metabolic activity. The metrics derived from this device would enhance the reproducibility and quality of target cell production.
{"title":"Compact lens-free imager using thin-film transistor for long-term quantitative monitoring of stem cell culture and cardiomyocyte production","authors":"Taishi Kakizuka, Tohru Natsume, Takeharu Nagai","doi":"10.1039/d4lc00528g","DOIUrl":"https://doi.org/10.1039/d4lc00528g","url":null,"abstract":"With advancements in human induced pluripotent stem cell (hiPSC) technology, there is an increasing demand for quality control techniques to manage the long-term process of target cell production effectively. While monitoring systems designed for use within incubators are promising for assessing culture quality, existing systems still face challenges in terms of compactness, throughput, and available metrics. To address these limitations, we have developed a compact and high-throughput lens-free imaging device named INSPCTOR. The device is as small as a standard culture plate, which allows for the installation of multiple units within an incubator. INSPCTOR utilises a large thin-film transistor image sensor, enabling simultaneous observation of six independent culture environments, each approximately 1 cm<small><sup>2</sup></small>. With this device, we successfully monitored the confluency of hiPSC cultures and identified the onset timing of epithelial-to-mesenchymal transition during mesodermal induction. Additionally, we quantified the beating frequency and conduction of hiPSC-derived cardiomyocytes by using high-speed imaging modes. This enabled us to identify the onset of spontaneous beating during differentiation and assess chronotropic responses in drug evaluations. Moreover, by tracking beating frequency over 10 days of cardiomyocyte maturation, we identified week-scale and daily-scale fluctuations, the latter of which correlated with cellular metabolic activity. The metrics derived from this device would enhance the reproducibility and quality of target cell production.","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":null,"pages":null},"PeriodicalIF":6.1,"publicationDate":"2024-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142431706","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}
Chunqiu Zhang, Ning Rong, Ziyi Lin, Pengqi Li, Jingyao Shi, Wei Zhou, Lili Niu, Fei Li, Rongxin Tang, Lei Li, Long Meng
Assisted reproductive technology (ART) has emerged as a crucial method in modern medicine for tackling infertility. However, the success of fertilization depends on the quality and quantity of sperm, often necessitating invasive surgical intervention, which presents challenges for non-invasive in vitro fertilization. Acoustic microfluidics technology has found widespread application across various biological contexts. In this paper, we propose to introduce a novel approach using asymmetric acoustic streaming generated by a single interdigital transducer (IDT) to enhance sperm concentration and improve fertilization in vitro, particularly in cases of moderate oligozoospermia. The concentration of particles increased approximately 6-fold in the central region after acoustic enrichment. Moreover, sperm motility was significantly improved without additional DNA fragmentation, and all the oocytes remained viable after 5 min of acoustic enrichment. Notably, acoustic enrichment accelerated fertilization and embryo development, leading to a higher fertilization rate and faster cleavage speed. Specifically, within 36 hours, the multiple-cell embryo ratio was significantly increased compared to the control group. This finding further validates the feasibility and non-invasiveness of acoustic enrichment for sperm fertilization in vitro. This work provides a promising tool for in vitro fertilization, holding significant implications for assisted reproduction.
辅助生殖技术（ART）已成为现代医学解决不孕不育问题的重要方法。然而，受精成功与否取决于精子的质量和数量，通常需要进行侵入性手术干预，这给无创体外受精带来了挑战。声学微流控技术已在各种生物领域得到广泛应用。在本文中，我们提出了一种新方法，利用单个趾间换能器（IDT）产生的不对称声流来提高精子浓度，改善体外受精，尤其是在中度少精症的情况下。声波富集后，中央区域的颗粒浓度增加了约 6 倍。此外，精子的运动能力也得到了明显改善，而且不会造成额外的 DNA 断裂，所有卵母细胞在声学富集 5 分钟后仍能存活。值得注意的是，声学富集加速了受精和胚胎发育，使受精率更高，裂殖速度更快。具体来说，与对照组相比，36 小时内多细胞胚胎比率明显增加。这一发现进一步验证了声学富集技术在体外精子受精方面的可行性和非侵入性。这项工作为体外受精提供了一种前景广阔的工具，对辅助生殖具有重要意义。
{"title":"Acoustic enrichment of sperm for in vitro fertilization","authors":"Chunqiu Zhang, Ning Rong, Ziyi Lin, Pengqi Li, Jingyao Shi, Wei Zhou, Lili Niu, Fei Li, Rongxin Tang, Lei Li, Long Meng","doi":"10.1039/d4lc00604f","DOIUrl":"https://doi.org/10.1039/d4lc00604f","url":null,"abstract":"Assisted reproductive technology (ART) has emerged as a crucial method in modern medicine for tackling infertility. However, the success of fertilization depends on the quality and quantity of sperm, often necessitating invasive surgical intervention, which presents challenges for non-invasive in vitro fertilization. Acoustic microfluidics technology has found widespread application across various biological contexts. In this paper, we propose to introduce a novel approach using asymmetric acoustic streaming generated by a single interdigital transducer (IDT) to enhance sperm concentration and improve fertilization in vitro, particularly in cases of moderate oligozoospermia. The concentration of particles increased approximately 6-fold in the central region after acoustic enrichment. Moreover, sperm motility was significantly improved without additional DNA fragmentation, and all the oocytes remained viable after 5 min of acoustic enrichment. Notably, acoustic enrichment accelerated fertilization and embryo development, leading to a higher fertilization rate and faster cleavage speed. Specifically, within 36 hours, the multiple-cell embryo ratio was significantly increased compared to the control group. This finding further validates the feasibility and non-invasiveness of acoustic enrichment for sperm fertilization in vitro. This work provides a promising tool for in vitro fertilization, holding significant implications for assisted reproduction.","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":null,"pages":null},"PeriodicalIF":6.1,"publicationDate":"2024-10-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142405505","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}
Shashwat Agarwal, Marcos Cortes-Medina, Jacob C. Holter, Alex Avendano, Joseph W. Tinapple, Joseph M. Barlage, Miles M. Menyhert, Lotanna M. Onua, Jonathan W. Song
Blood and lymphatic vessels in the body are central to molecular and cellular transport, tissue repair, and pathophysiology. Several approaches have been employed for engineering microfabricated blood and lymphatic vessels in vitro, yet these approaches invariably require specialized equipment, facilities, and research training beyond the capabilities of most biomedical laboratories. Here we present xurography as an inexpensive, accessible, and versatile rapid prototyping technique for engineering cylindrical and lumenized microvessels. Using a benchtop xurographer, or a cutting plotter, we fabricated modular multi-layer poly(dimethysiloxane) (PDMS) -based microphysiological systems (MPS) that house endothelial-lined microvessels approximately 260μm in diameter embedded within a user-defined 3-D extracellular matrix (ECM). We validated the vascularized MPS (or vessel-on-a-chip) by quantifying changes in blood vessel permeability due to the pro-angiogenic chemokine CXCL12. Moreover, we demonstrated the reconfigurable versatility of this approach by engineering three different vessel-ECM arrangements, which were obtained by minor adjustments to one or two steps of the fabrication process. Several of these arrangements, such as ones that incorporate close-ended vessel structures and spatially distinct ECM compartments along the same microvessel, cannot be readily achieved with other microfabrication strategies. Therefore, we anticipate that our low-cost and easy-to-implement fabrication approach will facilitate wider accessibility of MPS with tunable vascular architectures and ECM components while reducing the turnaround time required for iterative designs.
{"title":"Rapid low-cost assembly of modular microvessel-on-a-chip with benchtop xurography","authors":"Shashwat Agarwal, Marcos Cortes-Medina, Jacob C. Holter, Alex Avendano, Joseph W. Tinapple, Joseph M. Barlage, Miles M. Menyhert, Lotanna M. Onua, Jonathan W. Song","doi":"10.1039/d4lc00565a","DOIUrl":"https://doi.org/10.1039/d4lc00565a","url":null,"abstract":"Blood and lymphatic vessels in the body are central to molecular and cellular transport, tissue repair, and pathophysiology. Several approaches have been employed for engineering microfabricated blood and lymphatic vessels in vitro, yet these approaches invariably require specialized equipment, facilities, and research training beyond the capabilities of most biomedical laboratories. Here we present xurography as an inexpensive, accessible, and versatile rapid prototyping technique for engineering cylindrical and lumenized microvessels. Using a benchtop xurographer, or a cutting plotter, we fabricated modular multi-layer poly(dimethysiloxane) (PDMS) -based microphysiological systems (MPS) that house endothelial-lined microvessels approximately 260μm in diameter embedded within a user-defined 3-D extracellular matrix (ECM). We validated the vascularized MPS (or vessel-on-a-chip) by quantifying changes in blood vessel permeability due to the pro-angiogenic chemokine CXCL12. Moreover, we demonstrated the reconfigurable versatility of this approach by engineering three different vessel-ECM arrangements, which were obtained by minor adjustments to one or two steps of the fabrication process. Several of these arrangements, such as ones that incorporate close-ended vessel structures and spatially distinct ECM compartments along the same microvessel, cannot be readily achieved with other microfabrication strategies. Therefore, we anticipate that our low-cost and easy-to-implement fabrication approach will facilitate wider accessibility of MPS with tunable vascular architectures and ECM components while reducing the turnaround time required for iterative designs.","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":null,"pages":null},"PeriodicalIF":6.1,"publicationDate":"2024-10-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142383747","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}
Maria Cristina Ceccarelli, Marie Celine Lefevre, Attilio Marino, Francesca Pignatelli, Katarzyna Krukiewicz, Matteo Battaglini, Gianni Ciofani
A significant challenge in the treatment of central nervous system (CNS) disorders is represented by the presence of the blood-brain barrier (BBB), a highly selective membrane that regulates molecular transport and restricts the passage of pathogens and therapeutic compounds. Traditional in vivo models are constrained by high costs, lengthy experimental timelines, ethical concerns, and interspecies variations. To address these limitations, in vitro models, particularly microfluidic BBB-on-a-chip devices, have been developed. These advanced models aim to more accurately replicate human BBB conditions by incorporating human cells and physiological flow dynamics. In this framework, here we developed an innovative microfluidic system that integrates thin-film electrodes for non-invasive, real-time monitoring of BBB integrity using electrochemical impedance spectroscopy (EIS). EIS measurements showed frequency-dependent impedance changes, indicating BBB integrity and distinguishing well-formed from non-mature barriers. The data from EIS monitoring was confirmed by permeability assays performed with a fluorescence tracer. The model incorporates human endothelial cells in a vessel-like arrangement to mimic the vascular component and three-dimensional cell distribution of human astrocytes and microglia to simulate the parenchymal compartment. By modeling the BBB-on-a-chip with an equivalent circuit, a more accurate trans-endothelial electrical resistance (TEER) value was extracted. The device demonstrated successful BBB formation and maturation, confirmed through live/dead assays, immunofluorescence and permeability assays. Computational fluid dynamics (CFD) simulations confirmed that the device mimics in vivo shear stress conditions. Drug crossing assessment was performed with two chemotherapy drugs: doxorubicin, with a known poor BBB penetration, and temozolomide, conversely specific drug for CNS disorders and able to cross the BBB, to validate the model predictive capability for drug crossing behavior. The proposed sensorized microfluidic device represents a significant advancement in BBB modeling, offering a versatile platform for CNS drug development, disease modeling, and personalized medicine.
{"title":"Real-time monitoring of a 3D blood-brain barrier model maturation and integrity with a sensorized microfluidic device","authors":"Maria Cristina Ceccarelli, Marie Celine Lefevre, Attilio Marino, Francesca Pignatelli, Katarzyna Krukiewicz, Matteo Battaglini, Gianni Ciofani","doi":"10.1039/d4lc00633j","DOIUrl":"https://doi.org/10.1039/d4lc00633j","url":null,"abstract":"A significant challenge in the treatment of central nervous system (CNS) disorders is represented by the presence of the blood-brain barrier (BBB), a highly selective membrane that regulates molecular transport and restricts the passage of pathogens and therapeutic compounds. Traditional in vivo models are constrained by high costs, lengthy experimental timelines, ethical concerns, and interspecies variations. To address these limitations, in vitro models, particularly microfluidic BBB-on-a-chip devices, have been developed. These advanced models aim to more accurately replicate human BBB conditions by incorporating human cells and physiological flow dynamics. In this framework, here we developed an innovative microfluidic system that integrates thin-film electrodes for non-invasive, real-time monitoring of BBB integrity using electrochemical impedance spectroscopy (EIS). EIS measurements showed frequency-dependent impedance changes, indicating BBB integrity and distinguishing well-formed from non-mature barriers. The data from EIS monitoring was confirmed by permeability assays performed with a fluorescence tracer. The model incorporates human endothelial cells in a vessel-like arrangement to mimic the vascular component and three-dimensional cell distribution of human astrocytes and microglia to simulate the parenchymal compartment. By modeling the BBB-on-a-chip with an equivalent circuit, a more accurate trans-endothelial electrical resistance (TEER) value was extracted. The device demonstrated successful BBB formation and maturation, confirmed through live/dead assays, immunofluorescence and permeability assays. Computational fluid dynamics (CFD) simulations confirmed that the device mimics in vivo shear stress conditions. Drug crossing assessment was performed with two chemotherapy drugs: doxorubicin, with a known poor BBB penetration, and temozolomide, conversely specific drug for CNS disorders and able to cross the BBB, to validate the model predictive capability for drug crossing behavior. The proposed sensorized microfluidic device represents a significant advancement in BBB modeling, offering a versatile platform for CNS drug development, disease modeling, and personalized medicine.","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":null,"pages":null},"PeriodicalIF":6.1,"publicationDate":"2024-10-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142383958","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}
Anne-Gaëlle Bourdat, Remco den Dulk, Bastien Serrano, Gervais Clarebout, Jean Porcherot, Armelle Keiser, Nicolas Sarrut, François Boizot, Xavier Mermet, Raymond Charles, Manuel Alessio, Patricia Laurent, Myriam Cubizolles
Improving food safety is crucial in the contexte of “One Health” approach. To guarantee product quality and safety, food industry, having a very high turnover rate, needs short time-to-result analyses. Therefore, user-friendly systems at the point-of-need are necessary, presenting relevant analytical performances and fullfiling the current regulations. To answer these challenges, a microfluidic platform integrating sample preparation and subsequent multiplex qPCR detection has been developed for on-site testing. The system consists of a fully automated instrument driving a microfluidic cartridge dedicated to the detection of multiple allergens in complex food matrices. The first part of the microfluidic cartridge contains pumps, reservoirs, valves and a filter to achieve DNA extraction, concentration and purification. Multiplex qPCR detection is carried out in the second part of the cartridge including a negative control chamber and five chambers for target analyte detection. The in-house developed instrument contains all functions to autonomously drive the microfluidic cartridge: pneumatic control for fluid actuation, thermal control for qPCR amplification and an optical sytem using three fluorescent wavelengths for multiplex detection of the target analytes and controls. We demonstrate the simultaneous detection of four different allergens – gluten, sesame, soy and hazelnut – from various complex food matrices. The turn-around-time from sample to result is close to two hours and controls in place validate the obtained results. For gluten, a direct comparison with ELISA shows that the regulatory threshold of 20 ppm is comfortably fulfilled. Moreover, all results are in agreement with external laboratory analyses performed in parallel on the same samples. Our findings confirm that the system can be used safely on-site without risk for cross contamination between various samples to be analysed. In conclusion, our microfluidic platform offers a robust method for on-site allergen management.
{"title":"Integrated microfluidic platform for on-site qPCR analysis: food allergen detection from sample to result","authors":"Anne-Gaëlle Bourdat, Remco den Dulk, Bastien Serrano, Gervais Clarebout, Jean Porcherot, Armelle Keiser, Nicolas Sarrut, François Boizot, Xavier Mermet, Raymond Charles, Manuel Alessio, Patricia Laurent, Myriam Cubizolles","doi":"10.1039/d4lc00570h","DOIUrl":"https://doi.org/10.1039/d4lc00570h","url":null,"abstract":"Improving food safety is crucial in the contexte of “One Health” approach. To guarantee product quality and safety, food industry, having a very high turnover rate, needs short time-to-result analyses. Therefore, user-friendly systems at the point-of-need are necessary, presenting relevant analytical performances and fullfiling the current regulations. To answer these challenges, a microfluidic platform integrating sample preparation and subsequent multiplex qPCR detection has been developed for on-site testing. The system consists of a fully automated instrument driving a microfluidic cartridge dedicated to the detection of multiple allergens in complex food matrices. The first part of the microfluidic cartridge contains pumps, reservoirs, valves and a filter to achieve DNA extraction, concentration and purification. Multiplex qPCR detection is carried out in the second part of the cartridge including a negative control chamber and five chambers for target analyte detection. The in-house developed instrument contains all functions to autonomously drive the microfluidic cartridge: pneumatic control for fluid actuation, thermal control for qPCR amplification and an optical sytem using three fluorescent wavelengths for multiplex detection of the target analytes and controls. We demonstrate the simultaneous detection of four different allergens – gluten, sesame, soy and hazelnut – from various complex food matrices. The turn-around-time from sample to result is close to two hours and controls in place validate the obtained results. For gluten, a direct comparison with ELISA shows that the regulatory threshold of 20 ppm is comfortably fulfilled. Moreover, all results are in agreement with external laboratory analyses performed in parallel on the same samples. Our findings confirm that the system can be used safely on-site without risk for cross contamination between various samples to be analysed. In conclusion, our microfluidic platform offers a robust method for on-site allergen management.","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":null,"pages":null},"PeriodicalIF":6.1,"publicationDate":"2024-10-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142377439","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}
Xiang Ren, Ruyu Zhou, George Ronan, S. Gulberk Ozcebe, Jiaying Ji, Satyajyoti Senapati, Keith March, Eileen Handberg, David Anderson, Carl J. Pepine, Hsueh-Chia Chang, Fang Liu, Pinar Zorlutuna
Rapid diagnosis of acute myocardial infarction (AMI) is crucial for optimal patient management. Accurate diagnosis and time of onset of an acute event can influence treatment plans, such as percutaneous coronary intervention (PCI). PCI is most beneficial within 3 hours of AMI onset. MicroRNAs (miRNAs) are promising biomarkers, with potential of early AMI diagnosis, since they are released before cell death and subsequent release of larger molecules [e.g., cardiac troponins (cTn)], and have greater sensitivity and stability in plasma versus cTn regardless of timing of AMI onset. However, miRNA-based AMI diagnosis can result in false positives due to miRNA content overlap between AMI and stable coronary artery disease (CAD). Accordingly, we explored the possibility of using a miRNA profile, rather than a single miRNA, to distinguish between CAD and AMI, as well as different stages following AMI onset. First we screened a library of 800 miRNA using plasma samples from 4 patient cohorts; no known CAD, CAD, ST-segment elevation myocardial infarction (STEMI) and STEMI followed by PCI, using Nanostring miRNA profiling technology. From this screening, based on machine learning SCAD and Lasso algorithms, we identified 9 biomarkers (miR-200b, miR-543, miR-331, miR-3605, miR-301a, miR-18a, miR-423, miR-142, and miR-132) that were differentially expressed in CAD, STEMI and STEMI-PCI and explored them to identify a miRNA profile for rapid and accurate AMI diagnosis. These 9 miRNAs were selected as the most frequently identified targets by SCAD and Lasso, as indicated in the “drum-plot” model in machine learning approach. We used age-matched patient samples to validate selected 9 miRNA biomarkers using a multiplexed ion-exchange membrane-based miRNA sensor platform, which measures specific miRNAs, and cTn as a control, simultaneously as a point-of-care device. Findings from this study will inform timely and accurate diagnosis of AMI and its stages, which are essential for effective management and optimal patient outcomes.
{"title":"Towards Real-Time Myocardial Infarction Diagnosis: A Convergence of Machine Learning and Ion-Exchange Membrane Technologies Leveraging miRNA Signatures","authors":"Xiang Ren, Ruyu Zhou, George Ronan, S. Gulberk Ozcebe, Jiaying Ji, Satyajyoti Senapati, Keith March, Eileen Handberg, David Anderson, Carl J. Pepine, Hsueh-Chia Chang, Fang Liu, Pinar Zorlutuna","doi":"10.1039/d4lc00640b","DOIUrl":"https://doi.org/10.1039/d4lc00640b","url":null,"abstract":"Rapid diagnosis of acute myocardial infarction (AMI) is crucial for optimal patient management. Accurate diagnosis and time of onset of an acute event can influence treatment plans, such as percutaneous coronary intervention (PCI). PCI is most beneficial within 3 hours of AMI onset. MicroRNAs (miRNAs) are promising biomarkers, with potential of early AMI diagnosis, since they are released before cell death and subsequent release of larger molecules [e.g., cardiac troponins (cTn)], and have greater sensitivity and stability in plasma versus cTn regardless of timing of AMI onset. However, miRNA-based AMI diagnosis can result in false positives due to miRNA content overlap between AMI and stable coronary artery disease (CAD). Accordingly, we explored the possibility of using a miRNA profile, rather than a single miRNA, to distinguish between CAD and AMI, as well as different stages following AMI onset. First we screened a library of 800 miRNA using plasma samples from 4 patient cohorts; no known CAD, CAD, ST-segment elevation myocardial infarction (STEMI) and STEMI followed by PCI, using Nanostring miRNA profiling technology. From this screening, based on machine learning SCAD and Lasso algorithms, we identified 9 biomarkers (miR-200b, miR-543, miR-331, miR-3605, miR-301a, miR-18a, miR-423, miR-142, and miR-132) that were differentially expressed in CAD, STEMI and STEMI-PCI and explored them to identify a miRNA profile for rapid and accurate AMI diagnosis. These 9 miRNAs were selected as the most frequently identified targets by SCAD and Lasso, as indicated in the “drum-plot” model in machine learning approach. We used age-matched patient samples to validate selected 9 miRNA biomarkers using a multiplexed ion-exchange membrane-based miRNA sensor platform, which measures specific miRNAs, and cTn as a control, simultaneously as a point-of-care device. Findings from this study will inform timely and accurate diagnosis of AMI and its stages, which are essential for effective management and optimal patient outcomes.","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":null,"pages":null},"PeriodicalIF":6.1,"publicationDate":"2024-10-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142377440","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}
Haidong Feng, Georgios Katsikis, India Napier, Gong Du, Josh Lim, Joseph Doyle, Scott R. Manalis, Linda G Griffith
Egg (oocyte) vitrification is the dominant method for preserving fertility for women of reproductive age. However, the method is typically performed by hand, requiring precise (~0.1 to 10 µL) and time-sensitive (~1 sec) liquid exchange of cryoprotectants (CPA) around eggs as well as fine handling of eggs (~100 µm) for immersion into liquid nitrogen (LN2). Here, we developed a microfluidic platform for programmable vitrification. Our platform is based on a millimeter-sized hanging droplet inside which a given egg is suspended and subjected to liquid exchanges within seconds. After programmable exposures to CPA, the egg is extracted from the liquid-air interface of the droplet using a motorized fine-tip instrument and immersed into LN2 for vitrification. To benchmark our platform with the manual method, we vitrified over a hundred mouse eggs and found comparable percentages (~95%) for post-vitrification survivability. In addition, our platform performs real-time microscopy of the egg thereby enabling future studies where its morphology may be linked to functional outcomes. Our study contributes to the ongoing efforts to enhance the automation of embryology techniques towards broader applications in reproductive medicine both for clinical and research purposes.
{"title":"Microfluidic Hanging Droplet as a Programmable Platform for Mammalian Egg Vitrification","authors":"Haidong Feng, Georgios Katsikis, India Napier, Gong Du, Josh Lim, Joseph Doyle, Scott R. Manalis, Linda G Griffith","doi":"10.1039/d4lc00428k","DOIUrl":"https://doi.org/10.1039/d4lc00428k","url":null,"abstract":"Egg (oocyte) vitrification is the dominant method for preserving fertility for women of reproductive age. However, the method is typically performed by hand, requiring precise (~0.1 to 10 µL) and time-sensitive (~1 sec) liquid exchange of cryoprotectants (CPA) around eggs as well as fine handling of eggs (~100 µm) for immersion into liquid nitrogen (LN2). Here, we developed a microfluidic platform for programmable vitrification. Our platform is based on a millimeter-sized hanging droplet inside which a given egg is suspended and subjected to liquid exchanges within seconds. After programmable exposures to CPA, the egg is extracted from the liquid-air interface of the droplet using a motorized fine-tip instrument and immersed into LN2 for vitrification. To benchmark our platform with the manual method, we vitrified over a hundred mouse eggs and found comparable percentages (~95%) for post-vitrification survivability. In addition, our platform performs real-time microscopy of the egg thereby enabling future studies where its morphology may be linked to functional outcomes. Our study contributes to the ongoing efforts to enhance the automation of embryology techniques towards broader applications in reproductive medicine both for clinical and research purposes.","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":null,"pages":null},"PeriodicalIF":6.1,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142377441","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}
Liver cancer is a significant global contributor to cancer-related mortality. Despite available targeted therapies, resistance to tyrosine kinase inhibitors (TKIs) like sorafenib and lenvatinib poses a formidable challenge. The tumor microenvironment (TME), inhabited by cancer-associated fibroblasts (CAFs), profoundly influences this resistance. To uncover the mechanisms, a 3D microfluidic chip replicating liver architecture was fabricated to probe the intricate mechanisms of TKI resistance. The chip design mirrors the hexagonal structure of liver lobules, situating liver cancer cells at the core, encircled by fibroblasts, with rigorous assessments confirming biocompatibility and consistent cell growth. After determining the IC50 values of sorafenib and lenvatinib in 2D co-culture, a transwell setup revealed drug resistance development in co-cultured cells. Within the 3D microfluidic chip, live/dead assays highlighted elevated viability under drug exposure, emphasizing fibroblast-driven drug resistance. The study identifies AHSG and CLEC3B as potential mediators of drug resistance in co-culture, significantly upregulated in the co-cultured medium. Functional tests confirmed their roles, as introducing recombinant AHSG and CLEC3B enhanced liver cancer cell resistance to sorafenib and lenvatinib in both 2D and 3D scenarios. In conclusion, by replicating the complex TME using microfluidic technology, this study sheds light on the roles of AHSG and CLEC3B as well as possible approaches for improving the effectiveness of liver cancer treatment.
{"title":"Exploring cancer-associated fibroblast-induced resistance to tyrosine kinase inhibitors in hepatoma cells using a liver-on-a-chip model.","authors":"Madhu Shree Poddar, Yu-De Chu, Gaurav Pendharkar, Cheng-Hsien Liu, Chau-Ting Yeh","doi":"10.1039/d4lc00624k","DOIUrl":"https://doi.org/10.1039/d4lc00624k","url":null,"abstract":"<p><p>Liver cancer is a significant global contributor to cancer-related mortality. Despite available targeted therapies, resistance to tyrosine kinase inhibitors (TKIs) like sorafenib and lenvatinib poses a formidable challenge. The tumor microenvironment (TME), inhabited by cancer-associated fibroblasts (CAFs), profoundly influences this resistance. To uncover the mechanisms, a 3D microfluidic chip replicating liver architecture was fabricated to probe the intricate mechanisms of TKI resistance. The chip design mirrors the hexagonal structure of liver lobules, situating liver cancer cells at the core, encircled by fibroblasts, with rigorous assessments confirming biocompatibility and consistent cell growth. After determining the IC<sub>50</sub> values of sorafenib and lenvatinib in 2D co-culture, a transwell setup revealed drug resistance development in co-cultured cells. Within the 3D microfluidic chip, live/dead assays highlighted elevated viability under drug exposure, emphasizing fibroblast-driven drug resistance. The study identifies AHSG and CLEC3B as potential mediators of drug resistance in co-culture, significantly upregulated in the co-cultured medium. Functional tests confirmed their roles, as introducing recombinant AHSG and CLEC3B enhanced liver cancer cell resistance to sorafenib and lenvatinib in both 2D and 3D scenarios. In conclusion, by replicating the complex TME using microfluidic technology, this study sheds light on the roles of AHSG and CLEC3B as well as possible approaches for improving the effectiveness of liver cancer treatment.</p>","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":null,"pages":null},"PeriodicalIF":6.1,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142360784","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}
JIJO Easo EASO GEORGE, Rajib Basak, Indresh Yadav, Chuan Jie Tan, Jeroen Anton van Kan, veronique arluison, Frank Wien, Johan RC van der Maarel
Regulation of protein mobility is a fundamental aspect of cellular processes. In this study, we examined the impact of DNA methylation on the diffusion of nucleoid associated protein Hfq. This protein is one of the most abundant proteins that shapes the bacterial chromosome and is involved in several aspects of nucleic acid metabolism. Fluorescence microscopy was employed to monitor the movement of Hfq along double-stranded DNA, which was stretched due to confinement within a nanofluidic channel device. The mobility of Hfq is significantly influenced by DNA methylation. Our results underscore the importance of bacterial epigenetic modifications in governing the movement of nucleoid associated proteins such as Hfq. Increased levels of methylation result in enhanced binding affinity, which in turn slows down the diffusion of Hfq on DNA. The reported control of protein mobility by DNA methylation has potential implications for the mechanisms involved in target DNA search processes and dynamic modelling of the bacterial chromosome.
调节蛋白质的流动性是细胞过程的一个基本方面。在这项研究中，我们研究了 DNA 甲基化对核仁相关蛋白 Hfq 扩散的影响。该蛋白质是塑造细菌染色体的最丰富的蛋白质之一，参与了核酸代谢的多个方面。研究人员利用荧光显微镜监测了 Hfq 沿着双链 DNA 的运动，双链 DNA 在纳米流体通道装置中由于受限而被拉伸。Hfq 的移动性受到 DNA 甲基化的显著影响。我们的研究结果凸显了细菌表观遗传修饰在调控核糖体相关蛋白（如Hfq）运动方面的重要性。甲基化水平的增加会增强结合亲和力，进而减缓 Hfq 在 DNA 上的扩散。据报道，DNA甲基化对蛋白质移动性的控制对目标DNA搜索过程和细菌染色体动态建模所涉及的机制具有潜在的影响。
{"title":"Effect of base methylation on binding and mobility of bacterial protein Hfq on double-stranded DNA","authors":"JIJO Easo EASO GEORGE, Rajib Basak, Indresh Yadav, Chuan Jie Tan, Jeroen Anton van Kan, veronique arluison, Frank Wien, Johan RC van der Maarel","doi":"10.1039/d4lc00628c","DOIUrl":"https://doi.org/10.1039/d4lc00628c","url":null,"abstract":"Regulation of protein mobility is a fundamental aspect of cellular processes. In this study, we examined the impact of DNA methylation on the diffusion of nucleoid associated protein Hfq. This protein is one of the most abundant proteins that shapes the bacterial chromosome and is involved in several aspects of nucleic acid metabolism. Fluorescence microscopy was employed to monitor the movement of Hfq along double-stranded DNA, which was stretched due to confinement within a nanofluidic channel device. The mobility of Hfq is significantly influenced by DNA methylation. Our results underscore the importance of bacterial epigenetic modifications in governing the movement of nucleoid associated proteins such as Hfq. Increased levels of methylation result in enhanced binding affinity, which in turn slows down the diffusion of Hfq on DNA. The reported control of protein mobility by DNA methylation has potential implications for the mechanisms involved in target DNA search processes and dynamic modelling of the bacterial chromosome.","PeriodicalId":85,"journal":{"name":"Lab on a Chip","volume":null,"pages":null},"PeriodicalIF":6.1,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142360543","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}
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