高密度微载体培养人肝癌细胞

High density Cultivalion of human Liver Cancer cell with microcarriers

目的研究人肝癌细胞在高密度微载体培养时的生长条件与培养方式,建立实验室规模高效、简便培养人肝癌细胞的方法。方法应用微载体CultiSpher-S和Cytodex-1在1 L SuperSpinner搅拌瓶中培养人肝癌细胞THCC-98,采用批式换液连续培养的方式,对细胞生长及营养物质葡萄糖利用、代谢产物乳酸生成进行分析。结果人肝癌细胞THCC-98在CultiSpher-S和Cytodex-1培养中葡萄糖日消耗量高达2 mmol/106细胞(2.17和2.3),乳酸日生成率为0.75～1.08 mmol/106细胞,最高浓度为9 mmol/L(9.2比9.5)。通过实施批式换液连续培养,换液0.3～0.9 V/d,可使细胞最高密度达(1.04比0.89)×107cells/ml。就两种微载体培养比较,CultiSpher-S由于具有更大的表面积/体积比,提供给细胞更大的生长空间,其所能达到的细胞密度高且营养物质利用更为有效。结论用1 L SuperSpinner搅拌瓶Cultisphere微载体批式换液连续高密度培养人肝癌细胞,细胞密度可达107cells/ml,批式收获细胞>1010。

Objective Explore the cell growth condition and cultivation mode of human liver cancer cell in high den- sity micromarrier culture and establish an efficient and compact culture method for human liver cancer cell culture. Methods Human Liver cancer cell THCC-98 has been cultured with microcarriers of CultiSpher - S and Cytodex - 1 in 1 L Super Spinner system using DMEM/high substrate glucose utilization and metabolite lactate glucose medium. Culture process was monitored by cell growth, production. Results In THCC-98 culture, glucose utilization rate reached as high as 2 mmol/106 cells per day （2.17 vs 2.3） ,Lactate production rate were 0.75 ~ 1.08 mmo//106 cells per day, the maximal concentration was 9 mmol/L （9.2 vs 9.5 ）. By performing batch medium exchange under continuous cultivation model at 0.3 ~ 0.9 work volume per day for 13 - 14 days, the maximal cell density were reached （ 1.04 vs 0.89 × 10^7 cells/ml）. By comparison, CuhiSpher - S has large surface/volume ratio, so it can reached higher cell density and more efficient nutrition utilization. Conclusion Using 1 L Super Spinner system with the batch medium exchange under continuous cultivation mode, human liver cancer cell density were reached 10^7 cells/ml and attained total cells 〉 10^10.