苹果浓缩汁中嗜酸耐热菌多克隆抗体的制备及纯化

Preparation and Purification of Polyclonal Antibody Against Aliyclobacillus acidoterrestris from Apple Juice Concentrate

从苹果浓缩汁中分离得到嗜酸耐热菌,首次以耐热菌繁殖体及芽胞体分别作为免疫抗原,采用耳静脉注射及肌肉注射2种途径,对8只日本大耳白兔进行免疫,分别获得多克隆抗体。使用试管凝集法分别检测得到的抗体效价及其特异性,并采用饱和硫酸铵沉淀法及DEAE离子交换法纯化多克隆抗体,最终经SDS-PAGE电泳进行鉴定。以耳静脉注射和肌肉注射为免疫途径,制备得到的抗体效价分别达到1∶2560和1∶640。耳静脉注射得到的抗体效价高且免疫周期短,因此,耳静脉注射途径为最佳免疫途径;抗体特异性及纯化效果好。首次建立了苹果浓缩汁中耐热菌的家兔免疫程序,获得了多克隆抗体。

In order to build the rapid detection method of A. acidoterrestris from the Apple Juice Concentrate （AJC） using the enzyme - linked immunoassay following, the polyclonal antibody against A. acidoterrestris was prepared in this paper firstly. We used the propagule and brood - gemma of the thermo tolerant bacteria as the immune antigen, which were separated from the AJC. Immune tests were performed on eight large ear white rabbits with ear vein intravenous and intramuscular injection way respectively and the polyclonal antibodies were obtained. Titers and specifieities of the antibodies were detected by tube agglutination test. The polyclonal antibodies were purified by saturated ammonium sulfate and DEAE ion exchange chromatography, and verified by SDS -PAGE electrophoresis. The titers of polyclonal antibody obtained with ear vein intravenous and intramuscular injection way were 1 ： 2 560 and 1 ： 640, respectively. The results showed the antibody obtained by ear vein intravenous injection had higher titer and shorter immunization period than that obtained by intramuscular injection. The specificity and purification effect of antibody were well. The immunization procedure of rabbit with A. acidoterrestris from the AJC was built and its polyclonal antibody was obtained for the first time.