Passive pumping for the parallel trapping of single neurons onto a microsieve electrode array

Abstract

Recent advances in brain-on-a-chip technology have led to the development of modified microelectrode arrays. Previously, the authors have contributed to this exciting field of neuroscience by demonstrating a fabrication process for producing microsieve chips that contain three-dimensional (3D) micropores at the electrodes [termed microsieve electrode arrays (μSEAs)]. This chip allows us to trap hundreds of single neuronal cells in parallel onto the electrodes [B. Schurink and R. Luttge, J. Vac. Sci. Technol., B 31, 06F903 (2013)]. However, trapping the neurons reproducibly under gentle, biocompatible conditions remains a challenge. The current setup involves the use of a hand-operated syringe that is connected to the back of the μSEA chip with a polydimethylsiloxane (PDMS) construct. This makes the capture process rather uncontrolled, which can lead to either cell damage by shear stress or the release of trapped neurons when unplugging the syringe and PDMS constructs. Although, the authors could achieve an efficient capture rate of single neurons within the 3D micropores (80%–90% filling efficiency), cell culture performance varied significantly. In this paper, the authors introduce a passive pumping mechanism for the parallel trapping of neurons onto the μSEA chip with the goal to improve its biological performance. This method uses the capillary pumping between two droplets (a “pumping droplet” on one side of the chip and a “reservoir droplet” on the other side) to create a stable and controllable flow. Due to simplification of the handling procedure, omitting the use of a syringe and additional connections to the μSEA chip, the set-up is compatible with real time microscopy techniques. Hence, the authors could use optical particle tracking to study the trapping process and record particle velocities by video imaging. Analyzing the particle velocities in the passive pumping regime, the authors can confirm a gentle uniform particle flow through the 3D micropores. The authors show that passive pumping particle velocity can be tightly controlled (from 5 to 7.5 to 10.4 μm/s) simply by changing the droplet volume of the pumping droplets from 20, 40, and 60 μl and keeping the reservoir drop constant (10 μl). The authors demonstrate that neuron capturing efficiency and reproducibility as well as neuronal network formation are greatly improved when using this passive pumping approach.

Nanofabricating neural networks: Strategies, advances, and challenges

“…In the outlook, in next generation microfluidic nervous system-on-chip, systems could be supported and tailored for their culture conditions by a scaffolding plate in a similar fashion as it has been theoretically suggested by us 81 as a single neuron detector readout by means of a microsieve as a disposable and pluggable culture plate. Previously, in our group, Frimat et al 82 demonstrated these types of microsieve-scaffolded neural networks by seeding SH-SY5Y cells via passive flows in parallel to the highly organized cell capturing sites that are easy to monitor either by an electrical sensor array or standard epifluorescent microscopy as a monolayer but still preserve some level of three-dimensionality of the cells in the RoIs. Hence, this type of a modular designed NoC system could further serve standardization and functionalization opportunities.…”

Gaining Micropattern Fidelity in an NOA81 Microsieve Laser Ablation Process

“…Originally, these microsieves were made with a high pattern fidelity by silicon micromachining, as we demonstrated to pair single cells to electrodes [ 22 ]. This technique was developed by Schurink et al with a high potency for cell capture yield and survival [ 23 ]. Additive manufacturing using photo-curable polymers is an alternative for photo-lithography processes and can directly yield a final product structure, which allows for the implementation of complex 3D-multilayer patterns in a cost-accessible manner [ 24 , 25 , 26 , 27 ].…”