A Paired Bead and Magnet Array for Molding Microwells with Variable Concave Geometries

A spheroid culture is a useful tool for understanding cellular behavior in that it provides an in vivo-like three-dimensional environment. Various ...

Lab-on-a-CD Platform for Generating Multicellular Three-dimensional Spheroids

A three-dimensional spheroid cell culture can obtain more useful results in cell experiments because it can better simulate cell microenvironments of the ...

chung-ang university

Chung-Ang University

Well-designed microfluidic platforms can be excellent tools to eliminate bottleneck problems or issues that have arisen in biological fields by providing unprecedented high-resolution control of mechanical and chemical microenvironments for cell culture. Among such microtechnologies, the precise generation of biochemical concentration gradients has been highly regarded in the biorelated scientific fields; even today, the principles and mechanisms for gradient generation continue to be refined, and the number of applications for this technique is growing. Here, we review the current status of the concentration gradient generation technologies achieved in various microplatforms and how they have been and will be applied to biological issues, particularly those that have arisen from cancer research, stem cell research, and tissue engineering. We also provide information about the advances and future challenges in the technological aspects of microscale concentration gradient generation.

Microplatforms for gradient field generation of various properties and biological applications.

Stem cells have infinite potential for regenerative therapy thanks to their advantageous ability which is differentiable to requisite cell types for recovery and self-renewal. The microsystem has been proved to be more helpful to stem cell studies compared to the traditional methods, relying on its advantageous feature of mimicking in vivo cellular environments as well as other profitable features such as minimum sample consumption for analysis and multiprocedures. A wide variety of microsystems were developed for stem cell studies; however, regenerative therapy-targeted applications of microtechnology should be more emphasized and gain more attractions since the regenerative therapy is one of ultimate goals of biologists and bioengineers. In this review, we introduce stem cell researches harnessing well-known microtechniques (microwell, micropattern, and microfluidic channel) in view point of physical principles and how these systems and principles have been implemented appropriately for characterizing stem cells and finding possible regenerative therapies. Biologists may gain information on the principles of microsystems to apply them to find solutions for their current challenges, and engineers may understand limitations of the conventional microsystems and find new chances for further developing practical microsystems. Through the well combination of engineers and biologists, the regenerative therapy-targeted stem cell researches harnessing microtechnology will find better suitable treatments for human disorders.

Features of Microsystems for Cultivation and Characterization of Stem Cells with the Aim of Regenerative Therapy.

Spheroid cell culture is very useful for further understanding cellular behavior including motility and biochemical reaction since it mimics three-dimensional (3D) in vivo organ tissue. Among previously proposed various methods for spheroid production, such as hanging drop and spinner flask, microwell is a recently developed method harnessing microtechnology to produce uniform-sized spheroids. Although soft-lithography has been popular for creating microwell arrays, a 3D spherical geometry has been regarded as difficult to fabricate using conventional methods, or often requires complex fabrication processes and expensive equipment. Here, we propose a new method for fabricating concave microwells for cell spheroid production and culture. To demonstrate this method, we fabricated a 30 × 30 microwell array in 3 × 3 cm plates, utilizing metal beads, a through-hole array, and an assembly of small magnets. The spherical metal beads were used as a mold for the microwell, naturally creating the desired 3D concave microwell geometry. One of the key ideas was to place and hold each metal bead in the designated through-hole using the small magnet array. We also performed computational simulation of the magnetostatic force to design and observe the magnetic force field in detail. In addition, to provide a practical demonstration of the proposed system in cell biology, we created and cultured adipose-derived stem cell spheroids for 14 days for chondrogenic differentiation. This method allows further variations in microwell geometry that will enhance the method's applicability as a helpful tool for various studies in cell biology, cancer research, and tissue engineering.

Magnetic force-assisted self-locking metallic bead array for fabrication of diverse concave microwell geometries.

The engineered three-dimensional (3-D) cell cultivation system for the production of multicellular spheroids has attracted considerable attention due to its improved in vivo relevance to cellular communications compared to the traditional two-dimensional (2-D) cell culture platform. The formation and maintenance of cell spheroids in healthy condition is the critical factor for tissue engineering applications such as the repair of damaged tissues, the development of organ replacement parts, and preclinical drug tests. However, culturing spheroids in conventional isolated single wells show limit ted yield and maintenance periods due to the lack of proper supplies of nutrition as well as intercellular chemical signaling. Here we develop the novel networked concave microwell arrays for effective construction of 3-D multi-cellular spheroids. The proposed method provides a suitable structure for the diffusion of oxygen, water-soluble nutrients, and cytokines for cell-cell interactions among the spheroids in neighboring microwells. We have further demonstrated in hepatocyte spheroids-cultured networked concave microwells showed enhanced cell viability and albumin secretion compared to the un-networked control group for two weeks. Our results reveal multi-cellular functionality could be tuned up by networking individual 3-D spheroids without supplying additional chemicals or biological supplements. We anticipate our result to be used in high-throughput cellular screening platforms to study cell-cell interactions in response to diverse chemical stimuli as well as development of in vivo mimicking customized 3-D tissue culture system.

Networked concave microwell arrays for constructing 3-D cell spheroids.

A cell spheroid culture has the benefit of simulating in vivo three-dimensional cell environments. Microwell systems have been developed to mass-produce large quantities of uniform spheroids, and are frequently used in research areas, such as cell biology, anticancer drug development, and regenerative therapy. Recently reported concave-bottomed microwell systems have delivered more benefits in producing spheroids of higher quality and facilitating more effective research. However, microwell fabrication methods are often complicated or expensive, and there are inherent limitations in the functions and characteristics of existing microwells. Therefore, further studies on concave microwell systems are required. In this study, we fabricate spherical microwells with funnel-shaped entrance structures for spheroid culture; the shape is an upside-down omega ([Formula: see text]), and is thus named 'Omega-well'. The Omega-well array is fabricated using the capillary action of liquid polymer on the pins of a computer central processing unit, which is accomplished without requiring expensive materials or difficult procedures. Various characteristic analyses are performed by experiments and computer simulation. It is demonstrated that cell loss is minimized during cell seeding, a produced spheroid does not easily escape, and that crosstalk between microwells is significantly reduced. The novel fabrication method and Omega-well platform proposed in this study are highly practical, and thus will be useful tools in biology and pharmaceutical labs.

Gi-Hun Lee

School of Mechanical Engineering,
College of Engineering,
Department of Mechanical Engineering,
Graduate School,
School of Mechanical Engineering, College of Engineering,
Department of Mechanical Engineering, Graduate School

chung-ang university

A Paired Bead and Magnet Array for Molding Microwells with Variable Concave Geometries

Lab-on-a-CD Platform for Generating Multicellular Three-dimensional Spheroids

Gi-Hun Lee

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}School of Mechanical Engineering,College of Engineering,Department of Mechanical Engineering,Graduate School,School of Mechanical Engineering, College of Engineering,Department of Mechanical Engineering, Graduate School

chung-ang university

A Paired Bead and Magnet Array for Molding Microwells with Variable Concave Geometries

Lab-on-a-CD Platform for Generating Multicellular Three-dimensional Spheroids

School of Mechanical Engineering,
College of Engineering,
Department of Mechanical Engineering,
Graduate School,
School of Mechanical Engineering, College of Engineering,
Department of Mechanical Engineering, Graduate School