【作者】 王曦； 李彩梅； 劳文燕； 张德有； 马锐； 许宁； 杨旭琴； 朱亭玉； 刘英微； 李津；

【Author】 WANG Xi1,LI Cai-mei(1,2),LAO Wen-yan2,ZHANG De-you1,MA Rui1, XU Ning2,YANG Xu-qin1,ZHU Ting-yu1,LIU Ying-wei1,LI Jin(1,3) (1.Vaccine Research DepartmentⅡ,Beijing Tiantan Biological Products Co.,Ltd.,Beijing 10024,China 2.National Vaccine & Serum Institute;3.Beijing Union University; 4.Microbiology Department,Peking University Health Science Center)

【机构】 北京天坛生物制品股份有限公司； 北京生物制品研究所； 北京联合大学北苑校区； 北京大学医学部基础医学院病原生物学系；

【摘要】 目的:比较分析球磨法、低温超高压破碎法和YeastBuster细胞裂解液法三种技术破碎重组汉逊酵母细胞的适用性。方法:用球磨法、低温超高压破碎法和YeastBuster细胞裂解液法分别破碎表达HBsAg的重组汉逊酵母细胞样品和重组酿酒酵母细胞对照样品,测定细胞破碎率、细胞破碎后蛋白和HBsAg释出量,比较三种技术破碎细胞的效果。结果:球磨破碎法会将球磨罐和磨球中杂质成分磨入样品中,氧化锆罐-氧化锆珠磨入样品的杂质较少,但细胞的破碎率不高(60%～70%)。低温超高压破碎法破碎效果较好(70%～90%)。细胞裂解液法破碎细胞效果不佳(约60%),且影响对释出蛋白和抗原的准确检测,并可能破坏HBsAg。结论:低温超高压破碎法更加适用破碎重组汉逊酵母细胞。更多还原

【Abstract】 Objective:To compare the cell rupture effects of recombinant yeast Hansenula polymorpha with beadmilling, low-temperature high-pressure homogenization and YeastBuster protein extraction agent treatment.Methods:The bead-mill,low-temperature high-pressure homogenizer and YeastBuster protein extraction agent were employed to rupture the HBsAg expressing recombinant Hansenula polymorpha cell samples and the control recombinant Saccharomyces cerevisiae cells samples.The rates of cells rupture were evaluated by assay of protein and HBsAg released.Results:The components in either jars or beads could be abraded into samples when rupturing yeast cells with bead-milling,while agate jars and zirconia beads produced fewer impurities,the rates of cells rupture were 60%- 70%.The cells rupture effects with low-temperature high-pressure homogenizer were satisfactory which rupture rates of cell were 70%- 90%.The YeastBuster protein extraction agent could break 60%yeast cells,but interfered the assays accuracies of protein and HBsAg,and might spoil HBsAg.Conclusion:Compared to bead-milling and YeastBuster protein extraction agent,lowtemperature high pressure homogenization is more suitable to rupture the recombinant Hansenula polymorpha cells.更多还原