PVDF-g-PNIPAAm温敏膜用于Hela细胞黏附生长与脱附  

Thermal-Sensitive PVDF-g-PNIPAAm Membranes as a Substrate for Hela Cell Adhesion and Detachment

作　　者：李香琴[1] 刘天庆[1] 刘小红[1] 

机构地区：[1]大连理工大学干细胞与组织工程实验室,辽宁大连116024

出　　处：《高校化学工程学报》2016年第1期112-120,共9页Journal of Chemical Engineering of Chinese Universities

基　　金：国家自然科学基金项目(31170945)

摘　　要：传统的酶解法用于细胞收获容易导致细胞膜蛋白损伤,致使细胞功能缺失,而具有温敏特性的材料表面用于细胞培养时,可采用降温脱附法收获细胞,从而有效维护细胞的功能。研究将Hela细胞分别接种于常规空白培养板、PVDF膜以及四种不同PVDF-g-PNIPAAm温敏膜上,考察Hela细胞的黏附生长与降温脱附行为。培养过程中,观察细胞生长形态,检测细胞生长活率;收获时,观察细胞在从不同基底表面降温脱附行为,并对降温脱附法和胰酶消化法收获的细胞分别进行死活染色,计算细胞活率。结果表明,Hela细胞在不同基底表面均能正常黏附、铺展与生长;在四种不同PVDF-g-PNIPAAm温敏膜(M21,M32,M11,M23)上,M32与M11较更适宜细胞黏附与生长;当温度降低时Hela细胞不能从常规空白培养板及PVDF膜上自动脱附,但均能从四种温敏膜上自动脱附,脱附速率为M11>M32>M21>M23;降温脱附法比胰酶消化法收获的细胞活率更高,分别为(95±3.47)%与(81±2.26)%。PVDF-g-PNIPAAm温敏膜用于Hela细胞培养与收获的研究,为下一步采用温敏性PVDF-g-PNIPAAm中空纤维膜生物反应器大规模扩增并无损伤收获细胞奠定了基础。The traditional proteases for cell recovery can result in cell membrane protein damage, and lead to cell dysfunction and diseases. Alternatively, low temperature cell detachment from some thermal-sensitive surfaces can effectively maintain normal cell physiological functions. Hela cells were cultured on conventional culture plate, PVDF membrane and thermal-sensitive surface of PVDF-g-PNIPAAm to investigate their adhesion, growth and cooling detaching behaviors. Cell morphology was observed and cell viability was detected during cultivation. Cell cooling detachment behaviors from different substrate surfaces were observed, and the harvested cells by cooling detachment and trypsinization were stained by live/dead fluorescent dyes with cell viability calculated. The results show that Hela cell could adhere, spread and grow normally on different substrate surfaces, M32 and M11 as thermal-sensitive surfaces were more suitable for Hela cell adhesion and growth. Hela cell do not detach from conventional culture plate or PVDF membrane as temperature decreases, but it detaches from the thermal-sensitive surfaces and the detachment rates are in the order of M11 〉M32 〉M21 〉M23. Cell viability by cooling detachment is higher than that by trypsinization, with values of（95±3.47）% and（81±2.26）%, respectively. The research indicates that cooling detachment has better performance than trypsinization, which may help for cell expansion and harvest in large scale without damage in PVDF-g-PNIPAAm temperature sensitive hollow fiber membrane bioreactors.

关 键 词：聚N-异丙基丙烯酰胺 聚偏氟乙烯 温敏表面 细胞培养 降温脱附 

分 类 号：Q813.1[生物学—生物工程]