Preparation,Modification And Application Of Gelatin Microcarriers In Cell Culture

In this study,gelatin was used as the material to prepare a polydopamine-modified gelatin microcarrier,and a new cell culture method of "microsphere +polypeptide"was proposed and preliminarily verified.The preparation conditions of gelatin microspheres were explored and optimized in terms of oil phase,emulsifier,temperature,rotational speed,amount of emulsifier and amount of cross-linking agent.In order to increase the surface area of gelatin microspheres,gelatin microspheres with different morphologies were prepared by three methods,and the characterization of gelatin microspheres was determined by scanning electron microscope(SEM),Malvern particle size analyzer,infrared spectroscopy and other instruments.Polydopaminemodified gelatin microspheres were successfully prepared with porous structures on their surfaces.The characterization of polydopamine-modified gelatin microspheres was determined by scanning electron microscopy(SEM),infrared spectroscopy and other methods,and the application of cell culture was preliminarily carried out.The main research results are as follows:(1)Preparation of a porous polydopamine-modified gelatin microsphere.Under the conditions of gelatin concentration 30%,oil-water ratio 3:10,emulsifier spanu80,emulsification time 20 min,and crosslinking agent 50%glutaraldehyde 1m L,gelatin microspheres with an average particle size of about 233 um were prepared.During the preparation of gelatin microspheres,oil phase,emulsifier,temperature,rotational speed,amount of emulsifier and amount of crosslinking agent will all affect the average particle size of the microspheres.Three methods were designed with ethanol,water and isopropanol reagents,washed three times with ethanol and water,fully swollen in water,and freeze-dried at-80 °C to obtain gelatin microspheres with a larger surface area and a porous structure.Polydopamine modification was performed on the surface of gelatin microspheres for subsequent cell culture.(2)A new culture method of"microspheres + polypeptides"is proposed,using peptides to support the microspheres,cells can be adsorbed on the surface of the microspheres and increase in value,and this new culture method is preliminarily verified.The concentration of 0.1%polypeptide material and 0.2% microspheres can be uniformly dispersed in the medium,and the polypeptide material can support the microspheres well and remain stable within5 days of culture.The microspheres were used for the adsorption of porcine muscle satellite cells,and the AM-PI stained cells were observed under a confocal microscope.In order to quantify the level of cell proliferation,the CCK-8 kit was used to treat the cells cultured in the new culture method of "microsphere + polypeptide".The absorbance measurement at 450 nm showed that within 5 days of culture,"PDA-2g/L-2h,PDA-2g/L-4h,PDA-6g/L-2h four groups of microspheres and peptides cocultured with the value-added level is far greater than the 2D culture level.Therefore,it is proved that the new type of "microspheres + polypeptide"The culture method is feasible in cell culture.