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Here a protocol is presented to build a fast and non-destructive system for measuring cell or nucleus compressibility based on acoustofluidic microdevice. Changes in mechanical properties of tumor cells after epithelial-mesenchymal transition or ionizing radiation were investigated, demonstrating the application prospect of this method in scientific research and clinical practice.
Cell mechanics play an important role in tumor metastasis, malignant transformation of cells, and radiosensitivity. During these processes, studying the mechanical properties of the cells is often challenging. Conventional measurement methods based on contact such as compression or stretching are prone to cause cell damage, affecting measurement accuracy and subsequent cell culture. Measurements in adherent state can also affect accuracy, especially after irradiation since ionizing radiation will flatten cells and enhance adhesion. Here, a cell mechanics measurement system based on acoustofluidic method has been developed. The cell compressibility can be obtained by recording the cell motion trajectory under the action of the acoustic force, which can realize fast and non-destructive measurement in suspended state. This paper reports in detail the protocols for chip design, sample preparation, trajectory recording, parameter extraction and analysis. The compressibility of different types of tumor cells was measured based on this method. Measurement of the compressibility of nucleus was also achieved by adjusting the resonance frequency of the piezoelectric ceramic and the width of the microchannel. Combined with the molecular level verification of immunofluorescence experiments, the cell compressibility before and after drug-induced epithelial to mesenchymal transition (EMT) were compared. Further, the change of cell compressibility after X-ray irradiation with different doses was revealed. The cell mechanics measurement method proposed in this paper is universal and flexible and has broad application prospects in scientific research and clinical practice.
Cell mechanical properties play an important role in tumor metastasis, malignant transformation of cells, and radiosensitivity1,2. To gain an in-depth understanding of the role of cell mechanical properties in the above process, accurate measurement of cellular mechanics is critical, and the measurement should not cause damage to the cells for subsequent culture and analysis. The measurement process should be as fast as possible, otherwise cell viability may be affected if cells are removed from the cultivation environment for a long time.
Existing cell mechanics measurement methods....
1. Fabricating and assembly of the acoustofluidic microdevice
Here, the work presented a protocol for the construction of a fast and non-destructive cell compressibility measuring system based on acoustofluidic microdevice and demonstrated its advantages for measuring cell and nucleus under different situations. Figure 1 shows the schematic of the microfluidic channel. The components and assembly of the acoustofluidic microdevice are shown in Figure 2. Figure 3 shows the setup of the measureme.......
Commonly used cell mechanics measurement methods are AFM, micropipette aspiration, microfluidics methods, parallel-plate technique, optical tweezers, optical stretcher, and acoustic methods20. Microfluidics methods can work with three approaches: micro-constriction, extensional flow, and shear flow. Among them, optical stretcher, optical tweezers, acoustic methods, extensional flow, and shear flow approaches are non-contact measurements. In contrast to contact measurements, non-contact measurement.......
This study was supported by the National Natural Science Foundation of China (Grant numbers 12075330 and U1932165) and the Natural Science Foundation of Guangdong Province, China (Grant number 2020A1515010270).
....Name | Company | Catalog Number | Comments |
0.25% trypsin(1x) | GIBCO | 15050-065 | |
502 glue | Evo-bond | cyanoacrylate glue | |
A549 | ATCC | CCL-185 | lung adenocarcinoma |
Cytonucleoprotein and cytoplasmic protein extraction kit | Beyotime | P0027 | Contains cytoplasmic protein extraction reagents A and B |
Dulbecco’s modified Eagle medium (DMEM) | corning | 10-013-CVRC | |
Fetal Bovine Srum(FBS) | AUSGENEX | FBS500-S | |
HCT116 | ATCC | CCL247 | colorectal carcinoma |
Heat-resistant glass | Pyrex | ||
Leibovitz’s L-15 medium | GIBCO | 11415-064 | |
MCF-7 | ATCC | HTB-22Â | breast Adenocarcinoma |
MDA-MB-231 | ATCC | HTB-26Â | breast Adenocarcinoma |
Minimum Essential Medium (MEM) | corning | 10-010-CV | |
Penicillin-Streptomycin | GIBCO | 15140-122 | |
Phosphate buffer | corning | 21-040-cvc | |
PMSF | Beyotime | ST506 | 100mM |
Polybead Polystyrene Red Dyed Microsphere | polysciences | 15714 | The diameter of microshpere is 6.00µm |
propidium iodide(PI) | Sigma-Aldrich | P4170 | |
SYLGARD 184Silicone ELASTOMER | Dow-Corning | 1673921 | Contains prepolymers and curing agents |
Trypan Blue | Beyotime | C0011 |
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