Sonnenschein B, Bisping W
Zentralbl Bakteriol Orig A. 1976 Mar;234(2):247-59.
In order to detect minimal amounts of Clostridium botulinum toxins in animal tissue or food specimens it is necessary to use an extraction method which results in concentration of the botulinal toxins. In the present examinations, artificially contaminated canned beans were used to develop a suitable procedure for extraction and concentration of botulinal toxins A-E. The procedure consisted of 4 steps: 1. Canned beans were diluted 1:2 with 0.1 m phosphate buffer pH 6.0. 2. The diluted material was homogenised with an "Ultra-Turrax" homogeniser for 20 sec. 3. The monogenised material was centrifuged at 4000 rpm for 30 min. 4. 15 ml of supernatant was concentrated using a "Millipore ultrafiltration chamber" (with a membrane capable of excluding all material with a molecular weight above 25,000). A pressure of 1.5 atmospheres was applied until the terminal volume was 0.5 ml. Following extraction and concentration, the samples were assayed for botulinal toxin in mice. Using this assay the concentration of the five toxins were shown to be as follows: Type A toxin: 19.0-fold toxin concentration Type B toxin: 14.8-fold toxin concentration Type C toxin: 20.6-fold toxin concentration Type D toxin: 28.2-fold toxin concentration Type E toxin: 112.2-fold toxin concentration
为了检测动物组织或食品样本中痕量的肉毒梭菌毒素，有必要采用一种能使肉毒毒素浓缩的提取方法。在本实验中，使用人工污染的罐装豆子来开发一种适用于提取和浓缩A - E型肉毒毒素的方法。该方法包括4个步骤：1. 将罐装豆子用0.1 m pH 6.0的磷酸盐缓冲液按1:2稀释。2. 用“Ultra - Turrax”匀浆器将稀释后的材料匀浆20秒。3. 将匀浆后的材料以4000转/分钟的转速离心30分钟。4. 用“密理博超滤室”（配备能截留所有分子量大于25,000物质的膜）浓缩15毫升上清液。施加1.5个大气压直至终体积为0.5毫升。提取和浓缩后，在小鼠身上检测样本中的肉毒毒素。使用该检测方法，五种毒素的浓缩倍数如下：A型毒素：19.0倍毒素浓缩倍数；B型毒素：14.8倍毒素浓缩倍数；C型毒素：20.6倍毒素浓缩倍数；D型毒素：28.2倍毒素浓缩倍数；E型毒素：112.2倍毒素浓缩倍数
