Simple And Low-cost Fabrication Technology Of Concave Microcavity For Cell Spheroid Cultures

Recent studies have shown that the curved microcavity structure is more effective in preparing high-quality multicellular spheroids and establishing vitro microstructure models.It has received extensive attention and applications in research fields such as tissue engineering,regenerative medicine,cancer research,and drug screening.However,the current processing technology of curved microcavity has certain limitations,either the preparation process is complicated,the expensive equipment is required,the process is cumbersome and time-consuming,or the reproducibility of microcavity fabrication is difficult to guarantee.Therefore,there is an urgent need to develop a simple,low-cost,and fast curved microcavity manufacturing process to promote research and development in the fields of tissue engineering,regenerative medicine,and drug screening.This thesis proposes a simple and low-cost preparation method of curved microcavities,which can realize the rapid,flexible and low-cost production of cell spheroid culture chips in ordinary laboratories,and is used for high-quality and highthroughput culture of cell spheroids.The method can flexibly realize the production of curved concave microcavities and spherical microcavities by simply combining numerical control engraving technology,caramel inverted mold reflow technology and heterogeneous filling inverted mold technology.The main research contents are as follows:(1)A method of making curved concave microcavities using Computer Numerical Control(CNC)engraving technology combined with caramel mold reflow technology is proposed.Firstly,the optimal process parameters for processing the original concave microcavity of the CNC engraving machine were established,and the effects of the caramel reflow temperature and time on the surface finish and curvature of the microcavity were explored.Then,the material and microcavity of the culture chip were determined through cell culture experiments.The cavity aspect ratio,and then the minimum distance between adjacent microcavities was studied to achieve highthroughput culture of cell spheres and improve cell seeding efficiency.Finally,the curved concave microcavity chip was combined with the injection pipe array,realizing the highefficiency inoculation of a small number of cell samples,and further improving the efficiency of cell inoculation while saving cell raw materials.(2)A method for fabricating curved spherical microcavities based on heterogeneous filling inverted mold technology is proposed.The selection of two filling materials was determined,and the most suitable caramel filling concentration was successfully explored.Under this concentration condition,spherical microcavity chips with different sizes were fabricated,and were used for cultivation.A dense and uniform sphere of cells was produced,confirming the applicability of the proposed method.This method makes up for the shortcomings of the concave microcavity,effectively avoids the phenomenon that cells are easy to escape in a shallower chamber,and is more conducive to long-term cell culture.Compared with the existing manufacturing methods,the curved microcavity molding technology proposed in this thesis has the advantages of simplicity,low cost,and rapid prototyping.It is expected to promote the development of cancer biology,tissue engineering,and regenerative medicine.