2022, Vol. 26 ›› Issue (21): 3312-3318

Preparation of protein chips for screening matrix for Muse cell culture based on fibrinogen and gelatin

Li Xiuya1, Sheng Yang1, Wang Shengyi2, Li Yue2, Zhang Ruzhi2, Pan Yan3, Sun Yixin1, Deng Linhong3, Mark Bradley4, Zhang Rong1     

Abstract: BACKGROUND: Stress-tolerant multilineage differentiated (Muse) cells have excellent medical potential for regenerative medicine. However, at present, Muse cells need to be cultured and proliferated in vitro by suspension of cell aggregation, which is not conducive to large-scale expansion and application. Protein is one of the important components of extracellular matrix. Different proteins play different roles in cell growth. Therefore, the high-throughput method can be used to rapidly screen protein matrix that is conducive to the adhesion and proliferation of Muse cells on a two-dimensional surface.
OBJECTIVE: To identify suitable protein substrates for surface adhesion and passaging of Muse cells.
METHODS: Bovine serum albumin, bovine blood fibrin, and gelatin were used as raw materials, and their solutions were printed on glass slides by an ink jet printer and mixed on site to prepare protein microarray chips. The optimal protein combination for the adhesion of Muse cells was identified by using high-throughput screening approach. The Muse cells were cultured and passaged on the identified protein substrates for characterization.
RESULTS AND CONCLUSION: (1) Two protein combinations were obtained by high-throughput screening, which were beneficial to the adhesion of primary Muse cells. They were bovine serum albumin/gelatin and bovine blood fibrinogen/gelatin, with combination ratios of 7/17 for both. (2) CCK-8 assay showed that Muse cells proliferated well in all groups. Among them, cells cultured on plates coated with bovine blood fibrinogen/gelatin proliferated well in 6 days. (3) By analyzing the immunofluorescence staining results of passages 1 and 3 of Muse cells cultured on the protein-coated plate, it was found that the combination of bovine blood fibrinogen and gelatin with a ratio of 7/17 in weight supported the adhesion and proliferation of Muse cells for at least 3 passages, maintaining good stem cell characteristics.
Key words: Muse cell, cell therapy, cell amplification, bovine serum albumin, bovine blood fibrinogen, gelatin, high-throughput screening, stem cell characteristics