β-酪蛋白的电泳分离及径向免疫扩散法的优化

S. Djebara, K. El, K. Kanoun, Benali Mohammed, Benmalek Narimene
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摘要

β-酪蛋白(β-CN)是一种主要的乳制品蛋白,在储存过程中容易优先降解。本研究旨在利用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)分离分离牛乳中β-酪蛋白,并优化Mancini径向免疫扩散等免疫化学技术,以监测该乳蛋白在储存过程中的演变。为此,进行了一系列理化和细菌学分析,结果显示符合国际标准。这使我们能够使用优质牛奶对蛋白质进行更详细的研究,特别是β-酪蛋白,它被选为牛奶蛋白质质量的标志。将混合乳样品中总酪蛋白分离后进行电泳分离。使用了丙烯酰胺和双丙烯酰胺的各种组合,并保留了8 g / 230 mg对应的组合。总酪蛋白沉积量为833µg,为动物免疫提供了足够的b-酪蛋白浓度(300µg)。30只Wistar大鼠接种免疫方案35 d。获得的抗血清用于优化Mancini技术,并在3 - 69 h/+4°C的不同储存时间检测样品中的b-酪蛋白。检测到b-酪蛋白含量从0.85 mg ml-1下降到0.32 mg ml-1。这些结果证明了b-酪蛋白在低温下的脆弱性,并为牛奶的来源及其在乳制品或奶酪生产中的应用提供了信息。此外,我们的研究证实了免疫化学技术(如Mancini试验)在测定β-酪蛋白中的有效性。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Electrophoretic isolation of β-casein and optimization of a radial immunodiffusion test for bovine milk quality control
Beta-casein (β-CN) is a major dairy protein subject to preferential degradation during storage. Our study aimed to isolate β-casein from bovine milk under dissociative conditions by the electrophoretic technique using Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE) and optimize an immunochemical technique such as Mancini radial immunodiffusion in order to monitor the evolution of this milk protein during storage. For this purpose, a series of physicochemical and bacteriological analyses revealed conformity with international standards. This allowed us to use quality milk for more detailed studies of proteins, in particular β-casein, which was selected as a marker of milk protein quality. The total casein of mixed milk samples was isolated and then submitted to electrophoretic separation. Various combinations of acrylamide and bisacrylamide were used and the one corresponding to the ratio 8 g / 230 mg was retained. A total casein deposit of 833 µg provided an adequate b-casein concentration (300 µg) for consistent animal immunization. Thirty Wistar rats were subjected to an immunization protocol for 35 days. The obtained antisera were used to optimize the Mancini technique and to assay b-casein in our samples at different storage times ranging from 3 h to 69 h/+4°C. A decrease in b-casein content from 0.85 mg ml-1 to 0.32 mg ml-1 was detected. These results demonstrate the vulnerability of b-casein at low temperatures and provide information on the origin of milk and its application for dairy or cheese production. In addition, our study confirms the usefulness of immunochemical techniques such as the Mancini test in the determination of β-casein.
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