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Review
. 2022 Jan 3;13(1):80.
doi: 10.3390/mi13010080.

Advances in Single-Cell Printing

Xiaohu Zhou  1 , Han Wu  1 , Haotian Wen  1 , Bo Zheng  1
Affiliations
Review

Advances in Single-Cell Printing

Xiaohu Zhou et al. Micromachines (Basel). .

Abstract

Single-cell analysis is becoming an indispensable tool in modern biological and medical research. Single-cell isolation is the key step for single-cell analysis. Single-cell printing shows several distinct advantages among the single-cell isolation techniques, such as precise deposition, high encapsulation efficiency, and easy recovery. Therefore, recent developments in single-cell printing have attracted extensive attention. We review herein the recently developed bioprinting strategies with single-cell resolution, with a special focus on inkjet-like single-cell printing. First, we discuss the common cell printing strategies and introduce several typical and advanced printing strategies. Then, we introduce several typical applications based on single-cell printing, from single-cell array screening and mass spectrometry-based single-cell analysis to three-dimensional tissue formation. In the last part, we discuss the pros and cons of the single-cell strategies and provide a brief outlook for single-cell printing.

Keywords: cell array; inkjet printing; microfluidics; screening; single-cell analysis.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
(a) Overall microcontact printing of adhesive patterns using InnoStamp 40. (b) Fluorescence images of PC3-GFP cells immobilized on fibronectin micropatterns of various shapes. Patterns are depicted in dashed lines. Scale bar: 40 µm. Reproduced with permission from [59]. (c) Schematic illustration of the fabrication of polydopamine patterns on CYTOP-coated glass surface by negative microcontact printing. (d) Confocal image of the patterned single mouse mesenchymal stem cell array on polydopamine patterned CYTOP surface. Reproduced with permission from [60].
Figure 2
Figure 2
Acoustic field-assisted single-cell printing: (a-i) The setup of the acoustic picolitre droplet generator. (a-ii) A single 3T3 fibroblast cell with cell tracker dye was printed on a glass substrate (from left to right: white field image, fluorescent image, and the overlap image). Reproduced with permission from [45]. (b-i) Schematic illustration of the planar surface acoustic wave generators. (b-ii) Numerical simulation results of the surface acoustic wave-based 3D acoustic tweezer. (b-iii) The single 3T3 mouse fibroblast can be precisely printed to the desired position, either on the substrate to form a linear cell array (red arrow) or on the top of another cell (blue arrow). (b-iv) HeLa S3 cells were printed on a substrate to form the pattern of letters: “3” “D” “A” “T”. Reproduced with permission from [46].
Figure 3
Figure 3
(a-i) The single-cell printer consists of a dispenser chip (1) mounted to an aluminum case with a piezo stack actuator, a substrate, or microplate (2) for single-cell printing, an illumination system (3), a CCD camera (4) for cell detection, and a reservoir (5) for cell suspension loading. (a-ii) The size of the ROI was determined by the volume of the printed droplet. (a-iii) A single HeLa cell array was printed on a substrate. Reproduced with permission from [40]. (b) Schematic illustration of the real-time cellular recognition-based single-cell printing system using the microfluidic dispenser chip and droplet generation. Reproduced with permission from [41].
Figure 4
Figure 4
(a-i) Schematic illustration of the mechanism of the impedance-based single-cell printing system. The impedance signals of the flow-through cells trigger the actuator to generate and dispense the droplets containing single cells. (a-ii) The time-lapse images show the progress of printing a single HeLa cell. Reproduced with permission from [42]. (b-i) Schematic illustration of the system and mechanism of the dual microvalves-based single-cell screening and printing. (b-ii) HUVECs were captured by the valves under the pressure of 0.8 atm. (b-iii) the size distribution of the HUVECs’ suspension. (b-iv) the size of HUVECs screened by the valves. (b-v) Cell viability with different conditions. Reproduced with permission from [99].
Figure 5
Figure 5
(a-i) Schematic diagram of the fluorescence-activated cell sorter-based printed droplet microfluidics. (a-ii) The intracellular calcium release assay was screened on a single PC3 prostate cancer cell array with different concentrations of KCl. (a-iii) Box plots show the results of the single-cell-based intracellular calcium release assay. Reproduced with permission from [93]. (b-i) Schematic diagram of the inkjet printing process to fabricate the single-cell microarray. (b-ii) ATP-induced proliferation experiment indicated that the multi-cell group had a higher proliferation rate than the single cell. Reproduced with permission from [89].
Figure 6
Figure 6
(a-i) Schematic illustration of the experiment setup: (a-ii) Detection of the single MCF cell labeled with Rhodamine 6G. Reproduced with permission from [113]. (b-i) Schematic illustration of the three-phase single-cell printing (TP-SCP) system. (b-ii) Schematic diagram of the microfluidic chip with the three functional zones. (b-iii–b-v) MS spectra of (b-iii) 4T1 single cells, (b-iv) 293 single cells and (b-v) A2780 single cells. (b-vi) The classification result of the four types of cells. Reproduced with permission from [110].
Figure 7
Figure 7
(a) Schematic illustration of the high-definition single-cell printing (HD-SCP) system. (b) Schematic illustration of controlled spheroid formation with HD-SCP. (c–e) The size of the spheroids can be precisely controlled by the initial number of the printed single cells. (f) Multicellular spheroids formed by printing multiple cell types with different cell ratios. (g) Schematic illustration of dynamically controlled spheroid formation with HS-SCP. (h) Bioprinting multicellular Janus spheroids. Scale bars: 100 μm for (c) upper, and 200 μm for (c) lower, (e,f,h). Reproduced with permission from [116].
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References

    1. Altschuler S.J., Wu L.F. Cellular Heterogeneity: Do Differences Make a Difference? Cell. 2010;141:559–563. doi: 10.1016/j.cell.2010.04.033. - DOI - PMC - PubMed
    1. Wen L., Tang F. Boosting the Power of Single-Cell Analysis. Nat. Biotechnol. 2018;36:408–409. doi: 10.1038/nbt.4131. - DOI - PubMed
    1. Sun J., Gao L., Wang L., Sun X. Recent Advances in Single-Cell Analysis: Encapsulation Materials, Analysis Methods and Integrative Platform for Microfluidic Technology. Talanta. 2021;234:122671. doi: 10.1016/j.talanta.2021.122671. - DOI - PubMed
    1. Wang D., Bodovitz S. Single Cell Analysis: The New Frontier in ‘Omics’. Trends Biotechnol. 2010;28:281. doi: 10.1016/j.tibtech.2010.03.002. - DOI - PMC - PubMed
    1. Longo S.K., Guo M.G., Ji A.L., Khavari P.A. Integrating Single-Cell and Spatial Transcriptomics to Elucidate Intercellular Tissue Dynamics. Nat. Rev. Genet. 2021;22:627–644. doi: 10.1038/s41576-021-00370-8. - DOI - PMC - PubMed

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