Method of producing clostridium botulinum toxin using media containing plant-derived components and flexible closed container
一种肉毒梭菌、密闭容器的技术,应用在生物化学设备和方法、酵素、细菌等方向,能够解决存在高危因素、高投资成本、大劳动力等问题
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example 1
[0049] Example 1: Validation of medium composition to produce high levels of Clostridium botulinum toxin type A expressed outside the cell
[0050] The fact that a large amount of botulinum toxin type A is expressed extracellularly in media free of animal-derived components was verified using various plant media.
[0051] The media used are shown in Table 1 below.
[0052]
[0053] [Table 1]
[0054] [sheet]
[0055]
[0056] In Table 1, products manufactured by Beckton Dickinson and Fluka were used as soytone and phytotryptone, respectively. In addition, products manufactured by Becton-Dickinson were used as plant peptone and yeast extract, and products manufactured by Merck were used as glucose. The medium components of Table 1 were dissolved in water for injection (WFI), and the pH of the resulting solution was adjusted to 7.2 using NaOH.
[0057] In addition, 1% by weight of N-Z amine A (Sigma (Sigma): animal origin), 2% by weight of Bacto peptone (Proteose Pep...
example 2
[0075] In addition to not using Cultibag while culturing TM This example was performed in the same manner as Example 1 except that shaking was performed.
[0076] As a result, there was no substantial difference between the amount of toxin produced and that of Example 1.
example 3
[0077] Example 3: Isolation of meat type A from culture supernatant using media containing plant-derived components Clostridium toxin
[0078] Clostridium botulinum was cultured using medium 5 of Example 1 or 2, and botulinum toxin type A was isolated from the supernatant from which the cells had been removed. The culture conditions and time and the cell removal process were the same as in Example 1.
[0079] 0.1 N sulfuric acid was added to the bag containing 20 L of the culture supernatant to adjust the pH of the resulting solution to be in the range of 3.0 to 4.5, and thus, precipitate the toxin protein. Perform this procedure in a disposable sample bag that does not come into contact with the outside.
[0080]Next, when the precipitation was completed, the resulting solution was filtered through a pre-sterilized disposable filtration system (disposable sheet 2000.2 μm filter, manufactured by Sartorius Corporation) that was not in contact with the outside to remove the...
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