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Cell spheroids have been considered one potential model in the field of biological applications. This article describes protocols for scalably generating cell spheroids using a 3D acoustic assembly device, which provides an efficient method for the robust and rapid fabrication of uniform cell spheroids.
Cell spheroids are promising three-dimensional (3D) models that have gained wide applications in many biological fields. This protocol presents a method for manufacturing high-quality and high-throughput cell spheroids using a 3D acoustic assembly device through maneuverable procedures. The acoustic assembly device consists of three lead zirconate titanate (PZT) transducers, each arranged in the X/Y/Z plane of a square polymethyl methacrylate (PMMA) chamber. This configuration enables the generation of a 3D dot-array pattern of levitated acoustic nodes (LANs) when three signals are applied. As a result, cells in the gelatin methacryloyl (GelMA) solution can be driven to the LANs, forming uniform cell aggregates in three dimensions. The GelMA solution is then UV-photocured and crosslinked to serve as a scaffold that supports the growth of cell aggregates. Finally, masses of matured spheroids are obtained and retrieved by subsequently dissolving the GelMA scaffolds under mild conditions. The proposed new 3D acoustic cell assembly device will enable the scale-up fabrication of cell spheroids, and even organoids, offering great potential technology in the biological field.
3D in vitro culture models, which provide more in vivo-like structural and morphological characteristics compared to conventional 2D culture models, have been recognized as promising systems in various biomedical applications such as tissue engineering, disease modeling, and drug screening1,2,3. As one type of 3D culture model, cell spheroids typically refer to cell aggregation, creating 3D spheroidal structures characterized by enhanced cell-cell and cell-matrix interactions4,5,
1. Fabrication of the 3D acoustic assembly device
This study designed a 3D acoustic assembly device for mass manufacturing of cell spheroids. The acoustic device comprised a square chamber with two PZT transducers attached to the X-plane and Y-plane on the outer surface of the chamber and one PZT transducer on the chamber's bottom (Figure 1A,B). Three output channels from two function generators were connected to three power amplifiers to generate three independently sinusoidal signals to actuate the PZT transducers (
Efficient and stable fabrication of cell spheroids with high throughput using technologies like the 3D acoustic assembly device holds great promise for advancing biomedical engineering and drug screening1,2,3. This approach simplifies the mass production of cell spheroids through straightforward procedures.
However, there are critical factors to consider when using this acoustic device. The creation o.......
Tis work was supported by the National Key Research and Development Program of China (2022YFA1104600), and the Zhejiang Provincial Natural Science Foundation of China (LQ23H160011).
....Name | Company | Catalog Number | Comments |
0.22-μm filter | Merck | SLGSM33SS | Used for GelMA solution sterilization |
35 mm-cell culture dish | Corning | 430165 | Used for culturing cells |
Confocal microscope | Nikon | A1RHD25 | Fluorescent cell observation |
DiO dye | Beyotime | C1038 | Dye used to stain cells |
DMEM | Gibco | 12430054 | Cell culture media |
FBS | Gibco | 10099141C | Cell culture media supplement |
Function generator | Rigol | DG5352 | For RF signal generation |
GelMA | Regenovo | none | Used to prepare bioink |
GelMA lysis buffer | EFL | EFL-GM-LS-001 | Used to dissolve GelMA scaffolds |
Inverted microscope | Nikon | Ti-U | Cell observation |
LAP | Sigma-Aldrich | 900889 | Used as photoinitiator |
Live-Dead kit | Beyotime | C2015M | Cell vability analysis |
PBS | Gibco | 10010002 | Used as buffer |
Penicillin-streptomycin | Gibco | 15070063 | Prevent cell culture contamination |
Power amplifer | Minicircuit | LCY-22+ | Increase the voltage amplitude of the RF signal |
PZT transducers | Yantai Xingzhiwen Trading Co.,Ltd. | PZT-41 | Functional units for acoustic assembly device |
T25 cell culture flask | Corning | 430639 | Used for culturing cells |
Trypan blue | Gibco | 15250061 | Cell counting |
Trypsin-EDTA | Gibco | 25200056 | Cell dissociation enzyme |
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ISSN 2578-2614
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