Yu Xiaoli, Jiang Yanping, Zhang Songsong, Wang Caihong, Wang Ruichong, Zhang Lanlan, Tao Siming, Cui Wen, Li Jiaxuan, Qiao Xinyuan
Heilongjiang Key Laboratory for Animal Disease Control and Pharmaceutical Development, Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northeast Agricultural University, Harbin, China.
Department for Radiological Protection, Heilongjiang Province Center for Disease Control and Prevention, Harbin, China.
Front Microbiol. 2023 May 9;14:1174737. doi: 10.3389/fmicb.2023.1174737. eCollection 2023.
Bovine parvovirus (BPV) is a pathogen responsible for respiratory and digestive tract symptoms in calves and abortion and stillbirth in pregnant cows. In this study, we developed a colloidal gold immunochromatographic (GICG) strip with an enhanced signal for detecting BPV according to the double-antibody sandwich principle and an enzyme-based signal amplification system to amplify the signal. This system utilizes horseradish peroxidase reacting with a substrate solution containing 3,3',5,5-tetramethylbenzidine and dextran sulfate to obtain insoluble blue products on the test and control lines. We optimized different reaction conditions, including the amount of monoclonal antibodies (mAbs), pH of the colloidal gold solution, coating solution, blocking solution, sample pad treatment solution, antibody concentration in the control line, and antibody concentration in the detection line. The sensitivity of the signal-enhanced GICG strip showed that the minimum amount for detecting BPV was 10 TCID, 10 times higher than that of the traditional GICG strip. The results of the specificity test showed that the signal-enhanced GICG strip had no cross-reactivity with BRV, BVDV, or BRSV. The results of the repeatability test showed that the coefficient of variation between and within batches was less than 5%, showing good repeatability. Moreover, for validation, PCR and the signal-enhanced GICG strip were used to detect 280 clinical bovine fecal samples. The concordance rate compared with PCR was 99.29%. Hence, the developed strip exhibited high sensitivity and specificity for the detection of BPV. Therefore, this strip could be a rapid, convenient, and effective method for the diagnosis of BPV infection in the field.
牛细小病毒（BPV）是一种可导致犊牛呼吸道和消化道症状以及怀孕母牛流产和死产的病原体。在本研究中，我们根据双抗体夹心原理和基于酶的信号放大系统开发了一种用于检测BPV的信号增强型胶体金免疫层析（GICG）试纸条，以放大信号。该系统利用辣根过氧化物酶与含有3,3',5,5-四甲基联苯胺和硫酸葡聚糖的底物溶液反应，在检测线和对照线上获得不溶性蓝色产物。我们优化了不同的反应条件，包括单克隆抗体（mAb）的用量、胶体金溶液的pH值、包被液、封闭液、样品垫处理液、对照线抗体浓度和检测线抗体浓度。信号增强型GICG试纸条的灵敏度表明，检测BPV的最低量为10 TCID，比传统GICG试纸条高10倍。特异性试验结果表明，信号增强型GICG试纸条与牛轮状病毒（BRV）、牛病毒性腹泻病毒（BVDV）或牛呼吸道合胞病毒（BRSV）无交叉反应。重复性试验结果表明，批次间和批次内的变异系数均小于5%，具有良好的重复性。此外，为了进行验证，使用PCR和信号增强型GICG试纸条检测了280份临床牛粪便样本。与PCR相比的符合率为99.29%。因此，所开发的试纸条对BPV检测具有高灵敏度和特异性。因此，该试纸条可成为现场诊断BPV感染的快速、便捷且有效的方法。
