Browsing by Author "Sell, Scott A."
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Item Insert-based Microfluidics for 3D Cell Culture with Analysis(Springer Berlin Heidelberg, 2018-03-14) Chen, Chengpeng; Townsend, Alexandra D.; Hayter, Elizabeth A.; Birk, Hannah M.; Sell, Scott A.; Martin, R. ScottWe present an insert-based approach to fabricate scalable and multiplexable microfluidic devices for 3D cell culture and integration with downstream detection modules. Laser-cut inserts with a layer of electrospun fibers are used as a scaffold for 3D cell culture, with the inserts being easily assembled in a 3D-printed fluidic device for flow-based studies. With this approach, the number and types of cells (on the inserts) in one fluidic device can be customized. Moreover, after an investigation (i.e., stimulation) under flowing conditions, the cell-laden inserts can be removed easily for subsequent studies including imaging and cell lysis. In this paper, we first discuss the fabrication of the device and characterization of the fibrous inserts. Two device designs containing two (channel width = 260 μm) and four (channel width = 180 μm) inserts, respectively, were used for different experiments in this study. Cell adhesion on the inserts with flowing media through the device was tested by culturing endothelial cells. Macrophages were cultured and stimulated under different conditions, the results of which indicate that the fibrous scaffolds under flow conditions result in dramatic effects on the amount and kinetics of TNF-α production (after LPS stimulation). Finally, we show that the cell module can be integrated with a downstream absorbance detection scheme. Overall, this technology represents a new and versatile way to culture cells in a more in vivo fashion for in vitro studies with online detection modules.Item Microchip-based 3D-Cell Culture Using Polymer Nanofibers Generated by Solution Blow Spinning(Royal Society of Chemistry, 2017-04-21) Chen, Chengpeng; Townsend, Alexandra D.; Sell, Scott A.; Martin, R. ScottPolymer nano/micro fibers have found many applications including 3D cell culture and the creation of wound dressings. The fibers can be produced by a variety of techniques that include electrospinning, the primary disadvantage of which include the requirement for a high voltage supply (which may cause issues such as polymer denaturation) and lack of portability. More recently, solution blow spinning, where a high velocity sheath gas is used instead of high voltage, has been used to generate polymer fibers. In this work, we used blow spinning to create nano/microfibers for microchip-based 3D cell culture. First, we thoroughly investigated fiber generation from a 3D printed gas sheath device using two polymers that are amenable to cell culture (polycaprolactone, PCL and polystyrene, PS) as well as the parameters that can affect PCL and PS fiber quality. Using the 3D printed sheath device, it was found that the pressure of the sheath N2 and the concentration of polymer solutions determine if fibers can be produced as well as the resulting fiber morphology. In addition, we showed how these fibers can be used for 3D cell culture by directly depositing PCL fibers in petri dishes and well plates. It is shown the fibers have good compatibility with RAW 264.7 macrophages and the PCL fiber scaffold can be as thick as 178 ± 14 μm. PCL fibers created from solution blow spinning (with the 3D printed sheath device) were then integrated with a microfluidic device for the first time to fabricate a 3D cell culture scaffold with a flow component. After culturing and stimulating macrophages on the fluidic device, it was found that the integrated 3D fibrous scaffold is a better mimic of the extracellular matrix (as opposed to a flat, 2D substrate), with enhanced nitrite accumulation (product of nitric oxide release) from macrophages stimulated with lipopolysaccharide. PS fibers were also made and integrated in a microfluidic device for 3D culture of endothelial cells, which stayed viable for at least 72 hours (48 hours under the flowing conditions). This approach will be useful for future studies involving more realistic microchip-based culture models for studying cell-to-cell communication.Item A review of electrospinning manipulation techniques to direct fiber deposition and maximize pore size(Walter de Gruyter GmbH, 2017-06-18) Feltz, Kevin P.; Growney Kalaf, Emily A.; Chen, Chengpeng; Martin, R. Scott; Sell, Scott A.Electrospinning has been widely accepted for several decades by the tissue engineering and regenerative medicine community as a technique for nanofiber production. Owing to the inherent flexibility of the electrospinning process, a number of techniques can be easily implemented to control fiber deposition (i.e. electric/magnetic field manipulation, use of alternating current, or air-based fiber focusing) and/or porosity (i.e. air impedance, sacrificial porogen/sacrificial fiber incorporation, cryo-electrospinning, or alternative techniques). The purpose of this review is to highlight some of the recent work using these techniques to create electrospun scaffolds appropriate for mimicking the structure of the native extracellular matrix, and to enhance the applicability of advanced electrospinning techniques in the field of tissue engineering.Item Use of Electrospinning and Dynamic Air Focusing to Create Three-Dimensional Cell Culture Scaffolds in Microfluidic Devices(Royal Society of Chemistry, 2016-07-04) Chen, Chengpeng; Mehl, Benjamin T.; Sell, Scott A.; Martin, R. ScottOrgans-on-a-chip has emerged as a powerful tool for pharmacological and physiological studies. A key part in the construction of such a model is the ability to pattern or culture cells in a biomimetic fashion. Most of the reported cells-on-a-chip models integrate cells on a flat surface, which does not accurately represent the extracellular matrix that they experience in vivo. Electrospinning, a technique used to generate sub-micron diameter polymer fibers, has been used as an in vitro cell culture substrate and for tissue engineering applications. Electrospinning of fibers directly into a fully sealed fluidic channel using a conventional setup has not been possible due to issues of confining the fibers into a discrete network. In this work, a dynamic focusing method was developed, with this approach enabling direct deposition of electrospun fibers into a fully sealed fluidic channel, to act as a matrix for cell culture and subsequent studies under continuous flowing conditions. Scanning electron microscopy of electrospun polycaprolactone fibers shows that this method enables the formation of fibrous layers on the inner wall of a 3D-printed fluidic device (mean fiber size = 1.6 ± 0.6 μm and average pore size = 113 ± 19 μm²). Cells were able to be cultured in this 3D scaffold without the addition of adhesion proteins. Media was pumped through the channel at high flow rates (up to 400 μL/min) during a dynamic cell culture process and both the fibers and the cells were found to be strongly adherent. A PDMS fluidic device was also prepared (from a 3D printed mold) and coated with polycaprolactone fibers. The PDMS device enables optical detection and confocal imaging of cultured cells on the fibers. Finally, macrophages were cultured in the devices to study how the fibrous scaffold can affect cell behavior. It was found that under lipopolysaccharide stimulation, macrophages cultured on PCL fibers inside of a channel secreted significantly more cytokines than those cultured on a thin layer of PCL in a channel or directly on the inner wall of a channel. Overall, this study represents a new approach and technique for in vitro cell studies, where electrospinning can be used to easily and quickly create 3D scaffolds that can improve the culture conditions in microfluidic devices.