CCU50计算方法.docVIP

CCID50 (cell culture infective dose 50%) 细胞培养半数感染量。 　　主要用于检测病毒滴度。将病毒按照倍比稀释或梯度稀释后接种细胞(每个稀释度接种4孔或6孔细胞)，观察细胞CPE,通过Reed-Muench法计算CCID50，得到病毒滴度。这种方法主要用于测定病毒毒力滴度用.有了这个标准，就可以用100*CCID50或200*CCID50等,往下做一系列跟病毒有关的其他定量的实验,如中和实验中抗体效价测定等。更具体的操作方法计算和分析,参见病毒学书本，动物病毒学(殷震,刘景华主编)第二版上也有的。 S.O.P.2: ESTIMATION OF VIABLE MYCOPLASMA CONTENT OF CBPP VACCINES (Microtitration?Method) The test for the determination of viable Mycoplasma content of CBPP vaccines should be carried out on at least three vials of each freeze-dried or three bottles of each liquid vaccine batch. 针对于每批冻干或每批液体疫苗，CBPP疫苗中支原体活力测定实验应该至少测试三小瓶中。 The Test should be performed in parallel with a control vaccine preparation. For each test result to be valid the tire of the control vaccine preparation should not exceed 2 times the standard deviation of the calculated reference titre. Each titration shall be based on the use of ten-fold dilution series of the reconstituted vaccine and not less than 10 wells per dilution. 测试应该与疫苗准备同时进行。对于每个对照疫苗活力的结果不应该超过计算参考滴度标准差的2倍。每次滴定应该使用十倍稀释方法重组疫苗并且每次稀释不能少于10孔。 NB:?The minimum titre per (cattle) field dose should be at least 107?viable mycoplasmas. However taking into account the local storage and transportation conditions it is recommended that production laboratories should endeavour to deliver vaccine with at least 108?mycoplasmas per dose. NB：每段疫苗剂量的最低滴度至少应该在107，然而考虑到本地储存和运输条件，建议生产实验室应该努力每剂疫苗应该至少108. I. Equipment Sterile 96-well microtitration plates (Flat or U-bottomed) Plate sealer Glass of plastic bijoux bottltes (7 ml) Universal containers(30 ml) Pipetters. range 40 to 200 μl and 200 to 1000 μl Eppendorf multipette(R)?4780 or other type of suitable repetitive pipette Combitips, capacity: 2.5 ml, range 50–250 μl Microplate reader (viewer) 1.设备 无菌96孔微型滴定板（平板或U型底） 平板密封机 移液枪：40微升到200微升和200微升到1000微升 Eppendorf multipette(R)?移液枪或者其他类型适合的移液枪 枪头，规格2.5ml，50-250微升 酶标仪 II. Materials CBPP vaccine (freeze-dried or liquid) Water, pyrogen free, high purity, ionic strength less than 2.0 Siemens Co