Harshad Kamble

Microfluidic Technology for the Generation of Cell Spheroids and Their Applications

A three-dimensional (3D) tissue model has significant advantages over the conventional two-dimensional (2D) model. A 3D model mimics the relevant in-vivo physiological conditions, allowing a cell culture to serve as an effective tool for drug discovery, tissue engineering, and the investigation of disease pathology. The present reviews highlight the recent advances and the development of microfluidics based methods for the generation of cell spheroids. The paper emphasizes on the application of microfluidic technology for tissue engineering including the formation of multicellular spheroids (MCS). Further, the paper discusses the recent technical advances in the integration of microfluidic devices for MCS-based high-throughput drug screening. The review compares the various microfluidic techniques and finally provides a perspective for the future opportunities in this research area.

Single-Crystalline 3C-SiC anodically Bonded onto Glass: An Excellent Platform for High-Temperature Electronics and Bioapplications

Single-crystal cubic silicon carbide has attracted great attention for MEMS and electronic devices. However, current leakage at the SiC/Si junction at high temperatures and visible-light absorption of the Si substrate are main obstacles hindering the use of the platform in a broad range of applications. To solve these bottlenecks, we present a new platform of single crystal SiC on an electrically insulating and transparent substrate using an anodic bonding process. The SiC thin film was prepared on a 150 mm Si with a surface roughness of 7 nm using LPCVD. The SiC/Si wafer was bonded to a glass substrate and then the Si layer was completely removed through wafer polishing and wet etching. The bonded SiC/glass samples show a sharp bonding interface of less than 15 nm characterized using deep profile X-ray photoelectron spectroscopy, a strong bonding strength of approximately 20 MPa measured from the pulling test, and relatively high optical transparency in the visible range. The transferred SiC film also exhibited good conductivity and a relatively high temperature coefficient of resistance varying from -12 000 to -20 000 ppm/K, which is desirable for thermal sensors. The biocompatibility of SiC/glass was also confirmed through mouse 3T3 fibroblasts cell-culturing experiments. Taking advantage of the superior electrical properties and biocompatibility of SiC, the developed SiC-on-glass platform offers unprecedented potentials for high-temperature electronics as well as bioapplications.

Liquid marbles as bioreactors for the study of three-dimensional cell interactions

Liquid marble as a bioreactor platform for cell-based studies has received significant attention, especially for developing 3D cell-based assays. This platform is particularly suitable for 3D in-vitro modeling of cell-cell interactions. For the first time, we demonstrated the interaction of olfactory ensheathing cells (OECs) with nerve debris and meningeal fibroblast using liquid marbles. As the transplantation of OECs can be used for repairing nerve injury, degenerating cell debris within the transplantation site can adversely affect the survival of transplanted OECs. In this paper, we used liquid marbles to mimic the hostile 3D environment to analyze the functional behavior of the cells and to form the basis for cell-based therapy. We show that OECs interact with debris and enhanced cellular aggregation to form a larger 3D spheroidal tissue. However, these spheroids indicated limitation in biological functions such as the inability of cells within the spheroids to migrate out and adherence to neighboring tissue by fusion. The coalescence of two liquid marbles allows for analyzing the interaction between two distinct cell types and their respective environment. We created a microenvironment consisting of 3D fibroblast spheroids and nerve debris and let it interact with OECs. We found that OECs initiate adherence with nerve debris in this 3D environment. The results suggest that liquid marbles are ideal for developing bioassays that could substantially contribute to therapeutic applications. Especially, insights for improving the survival and adherence of transplanted cells.

Liquid Marble as Bioreactor for Engineering Three-Dimensional Toroid Tissues

Liquid marble is a liquid droplet coated with hydrophobic powder that can be used as a bioreactor. This paper reports the three-dimensional self-assembly and culture of a cell toroid in a slow-releasing, non-adhesive and evaporation-reducing bioreactor platform based on a liquid marble. The bioreactor is constructed by embedding a hydrogel sphere containing growth factor into a liquid marble filled with a suspension of dissociated cells. The hydrogel maintains the water content and concurrently acts as a slow-release carrier. The concentration gradient of growth factor induces cell migration and assembly into toroidal aggregates. An optimum cell concentration resulted in the toroidal (doughnut-like) tissue after 12 hours. The harvested cell toroids showed rapid closure of the inner opening when treated with the growth factor. We also present a geometric growth model to describe the shape of the toroidal tissue over time. In analogy to the classical two-dimensional scratch assay, we propose that the cell toroids reported here open up new possibilities to screen drugs affecting cell migration in three dimensions.

Superior Robust Ultrathin Single-Crystalline Silicon Carbide Membrane as a Versatile Platform for Biological Applications

Micromachined membranes are promising platforms for cell culture thanks to their miniaturization and integration capabilities. Possessing chemical inertness, biocompatibility, and integration, silicon carbide (SiC) membranes have attracted great interest toward biological applications. In this paper, we present the batch fabrication, mechanical characterizations, and cell culture demonstration of robust ultrathin epitaxial deposited SiC membranes. The as-fabricated ultrathin SiC membranes, with an ultrahigh aspect ratio (length/thickness) of up to 20 000, possess high a fracture strength up to 2.95 GPa and deformation up to 50 μm. A high optical transmittance of above 80% at visible wavelengths was obtained for 50 nm membranes. The as-fabricated membranes were experimentally demonstrated as an excellent substrate platform for bio-MEMS/NEMS cell culture with the cell viability rate of more than 92% after 72 h. The ultrathin SiC membrane is promising for in vitro observations/imaging of bio-objects with an extremely short optical access.

Magnetofluidic micromixer based on a complex rotating magnetic field

Rapid and efficient mixing of particles and fluids in a microfluidic system is of great interest for chemical and biochemical analysis. The present paper investigates magnetofluidic mixing induced by a rotating magnetic field from a number of permanent magnets. Numerical simulation shows the complex magnetic field in the mixing chamber. Simulated particle tracing predicts the trajectories of diamagnetic particles in a paramagnetic medium for the different stationary positions of the magnets. The experimentally obtained trajectories show negative magnetophoresis similar to that predicted by the simulation. However, the static configuration of the magnets cannot achieve mixing of the diamagnetic particles. We demonstrated that a rotating magnetic field could yield up to 86% mixing efficiency at a flow rate of 60 μL min−1 using a diluted ferrofluid of only 1% volume concentration.

An Electromagnetically Actuated Double-Sided Cell-Stretching Device for Mechanobiology Research

Cellular response to mechanical stimuli is an integral part of cell homeostasis. The interaction of the extracellular matrix with the mechanical stress plays an important role in cytoskeleton organisation and cell alignment. Insights from the response can be utilised to develop cell culture methods that achieve predefined cell patterns, which are critical for tissue remodelling and cell therapy. We report the working principle, design, simulation, and characterisation of a novel electromagnetic cell stretching platform based on the double-sided axial stretching approach. The device is capable of introducing a cyclic and static strain pattern on a cell culture. The platform was tested with fibroblasts. The experimental results are consistent with the previously reported cytoskeleton reorganisation and cell reorientation induced by strain. Our observations suggest that the cell orientation is highly influenced by external mechanical cues. Cells reorganise their cytoskeletons to avoid external strain and to maintain intact extracellular matrix arrangements.