Uniform cell seeding and generation of overlapping gradient profiles in a multiplexed microchamber device with normally-closed valves

Abstract

Generation of stable soluble-factor gradients in microfluidic devices enables studies of various cellular events such as chemotaxis and differentiation. However, many gradient devices directly expose cells to constant fluid flow and that can induce undesired responses from cells due to shear stress and/or wash out of cell-secreted molecules. Although there have been devices with flow-free gradients, they typically generate only a single condition and/or have a decaying gradient profile that does not accommodate long-term experiments. Here we describe a microdevice that generates several chemical gradient conditions on a single platform in flow-free microchambers which facilitates steady-state gradient profiles. The device contains embedded normally-closed valves that enable fast and uniform seeding of cells to all microchambers simultaneously. A network of microchannels distributes desired solutions from easy-access open reservoirs to a single output port, enabling a simple setup for inducing flow in the device. Embedded porous filters, sandwiched between the microchannel networks and cell microchambers, enable diffusion of biomolecules but inhibit any bulk flow over the cells.

In vitro quantitative analysis of Salmonella typhimurium preference for amino acids secreted by human breast tumor

“…The most critical drawback of hydrogel member based system, however, is that the hydrogels are not robust without an aqueous environment and the volume of the hydrogels can be changed significantly depending on the amount of the water absorption [14]. Moreover, the diffusion-based microfluidic devices which can generate the multiple chemical gradients have been developed [18][19][20][21]. Nevertheless, these devices have some drawbacks as comparing with the proposed device.…”

“…Nevertheless, these devices have some drawbacks as comparing with the proposed device. In some cases, the devices were constructed with complex multilayered microfluidic parts so that it is complicated to fabricate the devices [18][19][20]. In other case, it is not easy to realize the spatio-temporal control of the chemical gradient direction [21].…”

Polyester μ-assay chip for stem cell studies

“…15,38 In the first case, the goal is to create a highly controlled stem cell microenvironment and conduct high-throughput screening of differentiation cues. Thus, microfluidic valves and multiplexers are often incorporated into the devices for gradient and flow control, as shown by Mosadegh et al 39 and Gomez-Sjoberg et al, 40 which requires a high level of technical skill and potentially a programmable interface for automated operation. Similarly, Kim et al 41 enabled controlled on-chip cell loading, delivery of a range of chemical growth factors and immunostaining at the expense of a complicated 3-layer master mold fabrication, and the inclusion of externally driven microvalves.…”

A microfluidic co-culture system to monitor tumor-stromal interactions on a chip

“…18 Compared to conventional methods, the miniature size of microfluidic devices helps to create a microenvironment that better mimics the natural cellular environment. 19 In addition, the ability to precisely control flows 20 and generation of biochemical gradient 21,22 also facilitates to adjust and maintain favourable conditions for cell growth and introduce biochemical cues to influence the cell fate. 23 Researchers have taken advantage of these microfluidic platforms and successfully developed in vitro models for studying various important physiological phenomena, such as wound healing, 24,25 cancer metastasis, 26,27 and vascular angiogenesis.…”