Zhou Xiao-bo, Gao Jian-mei, Gao Chen, Zhang Bao-yun, Yuan Yu-kang, Dong Xiao-ping
Institute for Viral Disease Control and Prevention, China CDC, Beijing 100052, China.
Zhonghua Shi Yan He Lin Chuang Bing Du Xue Za Zhi. 2004 Jun;18(2):118-21.
To evaluate the influence ultrasonic processing on the aggregation of PrP(Sc) in the brain extracts prepared from the scrapie-infected hamsters, and to seek for the way to prepare lower molecular PrP(Sc) polymer.
The extracts of infected brains were prepared with a lysis solution, and treated with ultrasonics at various conditions during the different phases. The distribution and aggregation state of PrP(Sc) were analyzed by proteinase K treated. Western blot, and afterwards, quantitatively calculated with a commercially supplied software Image Totaltech.
The amount of PrP(Sc) in the supernatant of brain homogenates was 1.29-to 1.58-fold increased with appropriate sonication (15 s for 30 times). At the same conditions of ultrasound, the PrP amount in the supernatant prepared from the scrapie-infected hamster brain was significantly increased, whereas that prepared from healthy animal used as normal control showed little change. Comparative analyses of PrP(Sc) pellets prepared by high-speed centrifugation revealed that about 90% PrP(Sc) released into supernatant after ultrasound processing.
Appropriate sonication of homogenate of scrapie-infected brain increases the extracted amount of PrP(Sc), being favorable to laboratory diagnosis. Larger molecular PrP(Sc) aggregates can be crashed by ultrasonic processing, engendering lower molecular PrP(Sc) polymers.
评估超声处理对从感染羊瘙痒病的仓鼠制备的脑提取物中PrP(Sc)聚集的影响，并寻找制备低分子PrP(Sc)聚合物的方法。
用裂解液制备感染脑的提取物，并在不同阶段的各种条件下进行超声处理。用蛋白酶K处理、蛋白质印迹分析PrP(Sc)的分布和聚集状态，然后用商业供应的软件Image Totaltech进行定量计算。
适当超声处理（15秒，30次）后，脑匀浆上清液中PrP(Sc)的量增加了1.29至1.58倍。在相同的超声条件下，从感染羊瘙痒病的仓鼠脑制备的上清液中PrP的量显著增加，而作为正常对照的健康动物制备的上清液中PrP的量变化很小。对高速离心制备的PrP(Sc)沉淀进行比较分析发现，超声处理后约90%的PrP(Sc)释放到上清液中。
对感染羊瘙痒病的脑匀浆进行适当超声处理可增加PrP(Sc)的提取量，有利于实验室诊断。较大分子的PrP(Sc)聚集体可被超声处理破碎，产生低分子PrP(Sc)聚合物。
