An integrated microfluidic device for C. elegans early embryogenesis studies and drug assays
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The Caenorhabditis elegans embryo is a widely used model for the functional analysis of dynamic cellular processes, such as chromosome segregation, cytokinesis or lineage analysis. However, the conventional embryo preparation method that relies on manually dissecting gravid worms to extract embryos is somewhat time consuming and does not lend itself to high throughput assays. Here, we report a fully integrated microfluidic approach for C. elegans early embryogenesis assays with unprecedented accuracy and throughput. The device consists of a compressible microfluidic pillar-array chamber for robust and fast on-chip extraction of embryos from the uterus of gravid nematodes. Subsequently, embryos are immobilized by automated fluidic transfer in a microtrap array for individual tracking of a large number of embryos, including fragile mutants with drug-permeable eggshells. Our device allows high-resolution live imaging of very early events in embryogenesis, starting from the one-cell stage. We also demonstrate the feasibility of well-controlled compound application in versatile microfluidic pharmacological assays performed on early embryos.With the continuous development in nanoscience and nanotechnology, analytical techniques like surface-enhanced Raman spectroscopy (SERS) render structural and chemical information of a variety of analyte molecules in ultra-low concentration. Although this technique is making significant progress in various fields, the reproducibility of SERS measurements and sensitivity towards small molecules are still daunting challenges. In this regard, microfluidic surface-enhanced Raman spectroscopy (MF-SERS) is well on its way to join the toolbox of analytical chemists. This review article explains how MF-SERS is becoming a powerful tool in analytical chemistry. We critically present the developments in SERS substrates for microfluidic devices and how these substrates in microfluidic channels can improve the SERS sensitivity, reproducibility, and detection limit. We then introduce the building materials for microfluidic platforms and their types such as droplet, centrifugal, and digital microfluidics. Finally, we enumerate some challenges and future directions in microfluidic SERS. Overall, this article showcases the potential and versatility of microfluidic SERS in overcoming the inherent issues in the SERS technique and also discusses the advantage of adding SERS to the arsenal of microfluidics.
Surface-Enhanced Raman Spectroscopy
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In the budding yeast Saccharomyces cerevisiae, an actomyosin-based contractile ring is present during cytokinesis, as occurs in animal cells. However, the precise requirement for this structure during budding yeast cytokinesis has been controversial. Here we show that deletion of MYO1, the single myosin II gene, is lethal in a commonly used strain background. The terminal phenotype of myo1Δ is interconnected chains of cells, suggestive of a cytokinesis defect. To further investigate the role of Myo1p in cytokinesis, we conditionally disrupted Myo1 function by using either a dominant negative Myo1p construct or a strain where expression of Myo1p can be shut-off. Both ways of disruption of Myo1 function result in a failure in cytokinesis. Additionally, we show that amyo1Δ strain previously reported to grow nearly as well as the wild type contains a single genetic suppressor that alleviates the severe cytokinesis defects of myo1Δ. Using fluorescence time-lapse imaging and electron microscopy techniques, we show that cytokinesis in this strain is achieved through formation of multiple aberrant septa. Taken together, these results strongly suggest that the actomyosin ring is crucial for successful cytokinesis in budding yeast, but new cytokinetic mechanisms can evolve through genetic changes when myosin II function is impaired.
Septin
Budding
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Abstract Bio‐microfluidics applies biomaterials and biologically inspired structural designs (biomimetics) to microfluidic devices. Microfluidics, the techniques for constraining fluids on the micrometer and sub‐micrometer scale, offer applications ranging from lab‐on‐a‐chip to optofluidics. Despite this wealth of applications, the design of typical microfluidic devices imparts relatively simple, laminar behavior on fluids and is realized using materials and techniques from silicon planar fabrication. On the other hand, highly complex microfluidic behavior is commonplace in nature, where fluids with nonlinear rheology flow through chaotic vasculature composed from a range of biopolymers. In this Review, the current state of bio‐microfluidic materials, designs and applications are examined. Biopolymers enable bio‐microfluidic devices with versatile functionalization chemistries, flexibility in fabrication, and biocompatibility in vitro and in vivo. Polymeric materials such as alginate, collagen, chitosan, and silk are being explored as bulk and film materials for bio‐microfluidics. Hydrogels offer options for mechanically functional devices for microfluidic systems such as self‐regulating valves, microlens arrays and drug release systems, vital for integrated bio‐microfluidic devices. These devices including growth factor gradients to study cell responses, blood analysis, biomimetic capillary designs, and blood vessel tissue culture systems, as some recent examples of inroads in the field that should lead the way in a new generation of microfluidic devices for bio‐related needs and applications. Perhaps one of the most intriguing directions for the future will be fully implantable microfluidic devices that will also integrate with existing vasculature and slowly degrade to fully recapitulate native tissue structure and function, yet serve critical interim functions, such as tissue maintenance, drug release, mechanical support, and cell delivery.
Microtechnology
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Abstract Nanoparticles (NPs) have attracted tremendous interest in drug delivery in the past decades. Microfluidics offers a promising strategy for making NPs for drug delivery due to its capability in precisely controlling NP properties. The recent success of mRNA vaccines using microfluidics represents a big milestone for microfluidic NPs for pharmaceutical applications, and its rapid scaling up demonstrates the feasibility of using microfluidics for industrial‐scale manufacturing. This article provides a critical review of recent progress in microfluidic NPs for drug delivery. First, the synthesis of organic NPs using microfluidics focusing on typical microfluidic methods and their applications in making popular and clinically relevant NPs, such as liposomes, lipid NPs, and polymer NPs, as well as their synthesis mechanisms are summarized. Then, the microfluidic synthesis of several representative inorganic NPs (e.g., silica, metal, metal oxide, and quantum dots), and hybrid NPs is discussed. Lastly, the applications of microfluidic NPs for various drug delivery applications are presented.
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Integration of functional nanostructures within a microfluidic device can synergize the advantages of both unique properties of nanomaterials and diverse functionalities of microfluidics. In this paper, we report a novel and simple method for the in situ synthesis and integration of ZnO nanowires by controlled hydrothermal reaction within microfluidic devices. By modulating synthesis parameters such as the seed preparation, synthesis time, and heating locations, the morphology and location of synthesized nanowires can be easily controlled. The applications of such nanostructure-integrated microfluidics for particle trapping and chemiresistive pH sensing were demonstrated.
Nanomaterials
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This chapter summarizes the classical and recent achievements in the microstructured materials (MMs)/nanostructured materials (NMs) engineered from droplet microfluidics and their various applications. It overviews of MMs fabricated by droplet microfluidics, including the droplet formation mechanism and various microchips used to generate different droplets, the methods to prepare MMs templated from these droplets, and the unique and complex structures enabled by microfluidic techniques. The chapter presents basic synthesis methods for inorganic and organic NMs through droplet microfluidics, and the heterogeneous and multifunctional nanostructures from microfluidic platforms are also introduced. It emphasizes on the applications of the generated MMs/NMs, including drug delivery, cell encapsulation, TE, and analytical applications. The chapter discusses the current status and existing challenges and provides opinions on the directions of future development of droplet microfluidics in the synthesis of advanced MMs/NMs.
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Cytokinesis is the terminal step of the cell cycle during which a mother cell divides into daughter cells. Often, the machinery of cytokinesis is positioned in such a way that daughter cells are born roughly equal in size. However, in many specialized cell types or under certain environmental conditions, the cell division machinery is placed at nonmedial positions to produce daughter cells of different sizes and in many cases of different fates. Here we review the different mechanisms that position the division machinery in prokaryotic and eukaryotic cell types. We also describe cytokinesis-positioning mechanisms that are not adequately explained by studies in model organisms and model cell types.
Eukaryotic cell
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A molecular understanding of cytokinesis requires the detailed description of the protein complexes that perform central activities during this process. The proteins Hof1p, Cyk3p, Inn1p, and Myo1p each represent one of the four genetically defined and partially complementing pathways of cytokinesis in the yeast Saccharomyces cerevisiae. Here we show that the osmosensor Sho1p is required for correct cell-cell separation. Shortly before cytokinesis Sho1p sequentially assembles with Hof1p, Inn1p, and Cyk3p, into a complex (HICS-complex) that might help to connect the membrane with the actin-myosin ring. The HICS-complex is formed exclusively via the interactions between three SH3 domains located in Cyk3p, Hof1p, and Sho1p, and five acceptor sites found in Cyk3p, Hof1p, and Inn1p. Due to the overlapping binding specificities of its members the HICS-complex is best described as ensembles of isomeric interaction states that precisely coordinate the different functions of the interactors during cytokinesis.
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