摘要
目的:降低牛骨形态发生蛋白提取成本,增加提纯率,更加安全可靠地对其进行消毒贮存。方法:取1岁以内小牛长管状骨深低温保存、粉碎,依次以冰浴自来水、去郭子水洗涤,Urist非酶法分离提纯BMP,环氧乙烷熏蒸4℃低温消毒,SDS-聚丙烯酰胺凝胶电泳法测定提纯物蛋白质分子量,Wastar大鼠肌袋内植入BMP鉴定其生物活性。结果:用冰浴自来水部分地代替去离子水可节约后者用量且不影响BMP的提纯;在冰浴环境中缓慢搅拌0.6NHCI浸泡的骨粉并适时更换浸泡液可使脱钙更完全;将BMP以双层聚乙烯薄膜包裹后置于冷藏格内,适当延长环氧乙烷熏蒸消毒时间,既可达到确切的消毒效果,又不会影响BMP的生物活性。结论:采用改良理化法使脱钙更完全、可增加BMP提取量,低温环境下长时间熏蒸消毒不影响BMP的生物活性。
Objective： To search for a better method on purification and sterilization of bone morphognic protein （ BMP） from bovine bone. Methods- Cortical bone were obtained from steers under one- year- old, frozen in liquid N2 and washed in cold water for several times, then demineralized in HCI and defatted in chloroform methanol. Soluble proteins of BMP were extracted in CaCIe/NEM/PMSF and urea solution step by step. BMP was Sterilized and stored in a 4℃ circumstance. An analytical sodium dodecyl sulfate （ SDS） polyacrylamide gel electrophoresis was used for determination of proteins apparent molecular weights. The extracted BMP protein was implanted the thigh muscles of Waster strain mice to have a bioassay. Results： Using ice tap water but dehydronium water could save the latter and did not affect the extraction of BMP, demineralization of bone fraction could be more effective when it was stired in a cold circumstance in the 0.6N HCI solution. BMP was double wiapped at 4℃ could which be sterilizd for a longer time and would keep its bioactivety. Conclusions： Purification of BMP with improved Urist method weeds the demineralization of bone fraction more effective and could hervest more amount of product. BMP was double wiapped at 4 ℃ could be Sterilizd for a longer time and would keep its bioactivity.
出处
《华西医学》
CAS
2005年第4期702-703,共2页
West China Medical Journal
关键词
骨形态发生蛋白
提纯
消毒
生物活性
Bone morphogenetic protein （BMP）
Purification
Sterilization
Bioactivity
