布鲁氏菌血清IgM,IgG抗体胶体金免疫层析检测方法的建立及初步临床应用

Preliminary Clinical Application of Colloidal Gold Immunochromatographic Detection of Brucella IgM,IgG Antibodies

目的应用胶体金免疫层析技术建立一种快速检测布鲁氏菌IgM,IgG抗体的方法,并进行初步的性能验证。方法采用枸椽酸三钠还原法制备胶体金颗粒,标记纯化的抗重组布鲁氏菌抗原,将鼠抗人IgG抗体、鼠抗人IgM抗体和兔抗布鲁氏菌抗体喷在硝酸纤维素膜上作为检测线和质控线,对pH值和抗体浓度等条件优化;评估试剂盒灵敏度、特异度、交叉反应、准确度、精密度和干扰实验;用该方法对临床的400份血清进行检测,试管凝集试验验证其准确性。结果该试纸条最佳胶体金标记的pH值为7.5,标记重组抗原质量浓度为30μg/ml。方法的灵敏度和稳定性较好且与其他病原菌无交叉反应。血清总胆红素、血脂和血红蛋白浓度的增高会使试纸条显色结果减弱。对400份临床样本进行检测,其结果与试管凝集试验的符合率为93.75%。结论该试验建立的布鲁氏菌IgM/IgG抗体的胶体金免疫层析检测方法具有较好的特异度和稳定性,整个检测过程可在15min内完成,能够快速检测样品血清,适用于现场快速检测。

Objective A rapid detection method of IgM,IgG antibody against Brucella was established by colloidal gold immunochromatography,and the preliminary performance verification was carried out.Methods Colloidal gold particles were prepared by trisodium citrate reduction method,and the purified anti-recombinant Brucella antigen was labeled.A mouse antihuman IgG antibody,a mouse anti-human IgM antibody,and a rabbit anti-Brubella antibody were sprayed on a nitrocellulose membrane as a detection line and a quality control line,and optimized for pH,antibody concentration,and other conditions.Assessed the kit sensitivity,specificity,cross-reactivity,accuracy,precision,and interference experiments.This method was used to test 400 clinical serums,and the tube agglutination test verified its accuracy.Results The test strip had an optimum pH of 7.5 and a labeled recombinant antigen concentration of 30μg/ml.The method had good sensitivity and stability.It had no cross-reaction with other pathogens,and the increase of serum total bilirubin,blood lipid and hemoglobin concentration would weaken the coloration of test strips.For 400 clinical samples,the coincidence rate with the tube agglutination test was 93.75%.Conslusion The colloidal gold immunochromatographic assay for Brucella IgM,IgG antibody established in this experiment has good specificity and stability.The whole detection process can be completed within 15min,which can quickly detect sample serum and is suitable for rapid detection on site.