Inkjet-Printing Patterned Chip on Sticky Superhydrophobic Surface for High-Efficiency Single-Cell Array Trapping and Real-Time Observation of Cellular Apoptosis

Abstract

Single-cell assays have broad applications in cellular studies, tissue engineering, fundamental studies of cell-cell interactions, and understanding of cell-to-cell variations. Most existing methods for micron-sized cell patterning are still based on lithography-based microfabrication process. Thus, exploiting new mask-free strategies while maintaining high-precision single-cell patterning is still a great challenge. Here, we presented a facile, low-cost, and mask-free approach for constructing high-resolution patterning on sticky superhydrophobic (SH) substrates based on inkjet printing with ordinary precision. In this work, the SH surface with both high contact angle and relatively high contact angle hysteresis can not only obtain high-resolution spots but also avoid droplets bouncing behavior. We improved the feature size of printed protein spots as small as 4 μm, which is much smaller than protein spots used for single-cell trapping. Moreover, with the assistance of a narrow microchannel, the inkjet-printing patterned chip with fibronectin ink allows for fast and high-efficiency trapping of multiple single-cell arrays. Using this method, single-cell occupancy could reach approximately 81% within 30 min on subcellular-sized patterning chip, and there was no significant effect on cell viability. As a proof of concept, this chip has been applied to study the real-time apoptosis of single cells and demonstrated the potential in cells' heterogeneity analysis.

Micro‐/Nanostructured Interface for Liquid Manipulation and Its Applications

“…SPW had also been widely adopted in cell culture, analysis, [269,[283][284][285][286][287] and related drug screening. [288][289][290] Levkin group conducted many related researches.…”

Advances in Single-Cell Printing

“…Furthermore, single-cell bioprinting with the inkjet printing method could be achieved only by diluting the cell suspension to reduce the cell density [80,82,83]. Therefore, to increase the reliability and efficiency of single-cell printing, various new techniques have been developed to improve the dispensing efficiency of inkjet-like single-cell printing [40][41][42][44][45][46]68,[84][85][86][87][88][89][90][91][92][93][94][95][96][97][98][99].…”

“…As mentioned previously, single-cell analysis has become a powerful and indispensable tool in modern biological and medical research [2,3]. With the development of single-cell printing in the past decade, various single-cell printing-based single-cell analyses and applications have been performed, ranging from single-cell array-based screening [89,90,93,[107][108][109] and single-cell based mass spectroscopy [110][111][112][113][114][115] to live three-dimensional tissue formation [47,48,116].…”

Micro/Nanopatterned Superhydrophobic Surfaces Fabrication for Biomolecules and Biomaterials Manipulation and Analysis

“…The protocol was enough versatile to allow to switch the surface properties from completely superhydrophobic to superhydrophilic by just changing the process parameters. Combination of such approaches with surface patterning and controlled deposition of cells can also lead to a precise and controlled deposition of single cells or cells agglomerate [183]. The same group has proposed the use of SHSs as a platform to screen the biological performance of independent combinations of biomaterials, cells, and culture media [184,185], and as a mechanism for the formation of cell spheroids for their high-throughput parallel screening [186,187].…”