Altankov G, Brodvarova I, Rashkov I
Central Laboratory of Biophysics, Bulgarian Academy of Sciences, Sofia.
J Biomater Sci Polym Ed. 1991;2(2):81-9. doi: 10.1163/156856291x00089.
A novel technique for the synthesis of native collagen (type I)-coated gelatin microspheres was developed using the emulsion-polymerization principle, which is realized in four steps: emulsification, separation, stabilization, and protein modification. A 20% gelatin solution was emulsified in sunflower oil and the resulting beads were polymerized with glutaraldehyde. The novelty of our technique consists in the protein modification step, where we derivatized the beads by saturation of the free aldehyde groups on the cross-linked gelatin in the native collagen solution (0.3 mg/ml in 0.05 M Tris, pH 8.6) and further stabilized the beads with sodium borohydride. The resulting microspheres exhibited excellent properties as microcarriers for in vitro cell culturing using Vero cells as the model system. In comparison with pure gelatin beads, the cells attached more rapidly and grew faster on native collagen-coated beads. Approximately 90% of the Vero cells attached to the microcarrier after 1 h. After a lag period of nearly 24 h, the cells began to grow rapidly and reached confluence for 4-5 days, whereas the cells on pure gelatin beads reached the same density about 1 or 2 days later.
利用乳液聚合原理开发了一种合成天然胶原蛋白（I型）包被明胶微球的新技术，该技术通过四个步骤实现：乳化、分离、稳定化和蛋白质修饰。将20%的明胶溶液在向日葵油中乳化，所得微珠用戊二醛聚合。我们技术的新颖之处在于蛋白质修饰步骤，在该步骤中，我们通过用天然胶原蛋白溶液（在0.05 M Tris，pH 8.6中为0.3 mg/ml）饱和交联明胶上的游离醛基来衍生化微珠，并进一步用硼氢化钠稳定微珠。所得微球作为体外细胞培养的微载体表现出优异的性能，以Vero细胞作为模型系统。与纯明胶微珠相比，细胞在天然胶原蛋白包被的微珠上附着更快且生长更快。1小时后，约90%的Vero细胞附着在微载体上。经过近24小时的延迟期后，细胞开始快速生长并在4-5天达到汇合，而纯明胶微珠上的细胞约1或2天后达到相同密度。
