硝酸纤维素膜富集法检测志贺菌方法的建立及应用

Establishment and application of nitrocellulose membrane enrichment method for shigella detection

目的利用抗原抗体反应与培养法结合,建立先分离再增菌的增菌方法,提高志贺菌的培养检出率。方法将包被了志贺菌多价诊断血清的硝酸纤维素膜(NC膜)固定于拭子上,将拭子插入待测样本37℃孵育30 min,取出冲洗后,置新的营养肉汤管中再继续37℃增菌3 h,冲洗后将拭子涂在SS培养基平板上37℃培养18 h^24 h,计数阳性菌落数。结果通过对随机PCR法筛查阳性临床标本的检验,与传统方法相比较,用NC膜富集法增菌3 h,其检出志贺菌菌落的数目显著增高(t=2.56,P<0.05)。结论此方法大大提高了传统培养法的检出率以及增菌环节的选择性,减少了杂菌竟争性抑制和干扰。

Objective To establish a method to improve the detection rate of shigella based on combination of the antigen-antibody reaction and culture method,by which the shigella was firstly isolated and then cultured in medium for further analysis. Methods The nitrocellulose membrane was coated with antibody to shigella and fixed to a swab. Then the swab was inserted to the sample under test at 37 ℃ for 30min. After washing,the swab was inserted to a new nutrient broth at 37 ℃ for 3 h. The pure culture of shigella was done by washing swab stick with new medium and inoculated on SS medium plate for 18 h - 24 h at 37 ℃. The total number of shigella bacteria clones on SS medium plate was counted. Results Compared with traditional culture method,the number of shigella bacteria significantly increased with nitrocellulose membrane enrichment method（ t = 2. 56,P〈 0. 05）. Conclusion The nitrocellulose membrane enrichment method could greatly improve the detection rate of shigella bacteria and reduce the competitive suppression and interference of other bacteria to shigella growth.