Spatially selective cell treatment and collection for integrative drug testing using hydrodynamic flow focusing and shifting

Abstract

Hydrodynamic focusing capable of readily producing and controlling laminar flow facilitates drug treatment of cells in existing microfluidic culture devices. However, to expand applications of such devices to multiparameter drug testing, critical limitations in current hydrodynamic focusing microfluidics must be addressed. Here we describe hydrodynamic focusing and shifting as an advanced microfluidics tool for spatially selective drug delivery and integrative cell-based drug testing. We designed and fabricated a co-flow focusing, three-channel microfluidic device with a wide cell culture chamber. By controlling inlet flow rates of sample and two side solutions, we could generate hydrodynamic focusing and shifting that mediated precise regulation of the path and width of reagent and drug stream in the microfluidic device. We successfully validated a hydrodynamic focusing and shifting approach for spatially selective delivery of DiI, a lipophilic fluorophore, and doxorubicin, a chemotherapeutic agent, to tumor cells in our device. Moreover, subsequent flowing of a trypsin EDTA solution over the cells that were exposed to doxorubicin flow allowed us to selectively collect the treated cells. Our approach enabled downstream high-resolution microscopy of the cell suspension to confirm the nuclear delivery of doxorubicin into the tumor cells. In the device, we could also evaluate in situ the cytotoxic effect of doxorubicin to the tumor cells that were selectively treated by hydrodynamic flow focusing and shifting. These results show that hydrodynamic focusing and shifting enable a fast and robust approach to spatially treat and then collect cells in an optimized microfluidic device, offering an integrative assay tool for efficient drug screening and discovery.

Advances in high throughput cell culture technologies for therapeutic screening and biological discovery applications

“…Combined chemical and mechanical signal gradients on interspersed culture wells were designed to observe the effects of both drug concentration and shear stress on the viability of skin cancer cells and found that high shear stress has a synergistic effect with dosage on cell viability 68 . A co‐flow focusing, three‐channel microfluidic device was created that could direct path and width of a reagent or drug stream to spatially selective regions and the directed removal of only treated cells for further imaging and analysis 69 . A crossed laminar flow microfluidic system that could flow many different cell types to attach to printed microarray islands of Collagen I or antibodies that are then exposed to orthogonal streams of drug treatments enabled the HT assessment of drug and cell type interactions in a single experiment 70 …”

“…68 A co-flow focusing, three-channel microfluidic device was created that could direct path and width of a reagent or drug stream to spatially selective regions and the directed removal of only treated cells for further imaging and analysis. 69 A crossed laminar flow microfluidic system that could flow many different cell types to attach to printed microarray islands of Collagen I or antibodies that are then exposed to orthogonal streams of drug treatments enabled the HT assessment of drug and cell type interactions in a single experiment. 70 2.4 | Potential limitations of 2D and motivation to move to 3D HTS systems Two-dimensional HT culture systems offer robust screening of high numbers of soluble and insoluble effectors on cellular behavior and responses.…”