PDF(Pubmed)
Abstract：
Brucella BP26 proves to be a highly immunogenic antigen with excellent specificity in brucellosis detection. In China, the authorized use of the Bp26-deleted vaccine M5ΔBP26 for preventing small ruminant brucellosis highlights the importance of developing accurate detection methods targeting BP26, particularly for the diagnosis of differentiation between infected and vaccinated animals (DIVA). Using the traditional mouse hybridoma technique, we successfully obtained 12 monoclonal antibodies (mAbs) targeting BP26. The efficacy of these mAbs in detecting various animal brucellosis cases using the competitive ELISA method was evaluated. Among them, only the E10 mAb exhibited significant efficiency, being inhibited by 100, 97.62, and 100% of brucellosis-positive sera from cattle, small ruminants, and canines, respectively. The E10-based competitive enzyme-linked immunosorbent assay (cELISA) outperformed the BP26-based indirect enzyme-linked immunosorbent assay (iELISA) in accuracy, particularly for cattle and small ruminant brucellosis, with cELISA sensitivity reaching 97.62% compared to 64.29% for iELISA for small ruminants. Although cELISA showed slightly lower specificity than iELISA, it still maintained high accuracy in canine brucellosis detection. The epitope of mAb E10 was identified in the amino acid sequence QPIYVYPDDKNNLKEPTITGY, suggesting its potential as a diagnostic antigen for brucellosis. In conclusion, the E10-based cELISA presents an effective means of detecting animal brucellosis, particularly significant for DIVA diagnosis in China, where the BP26-mutant vaccine is widely used.
摘要：
布鲁氏菌BP26被证明是一种高度免疫原性的抗原，在布鲁氏菌病检测中具有出色的特异性。在中国,授权使用Bp26缺失疫苗M5ΔBP26预防小反刍动物布鲁氏菌病突出了开发针对BP26的准确检测方法的重要性，特别是对于诊断感染和接种疫苗的动物之间的区别（DIVA）。使用传统的小鼠杂交瘤技术，我们成功获得了12种靶向BP26的单克隆抗体。评估了这些mAb在使用竞争性ELISA方法检测各种动物布鲁氏菌病病例中的功效。其中,只有E10mAb表现出显著的效率，被牛的100、97.62和100％的布鲁氏菌病阳性血清抑制，小反刍动物,和犬科动物,分别。基于E10的竞争性酶联免疫吸附测定（cELISA）在准确性上优于基于BP26的间接酶联免疫吸附测定（iELISA），特别是对于牛和小反刍动物布鲁氏菌病，cELISA的灵敏度达到97.62%，而iELISA对小反刍动物的灵敏度为64.29%。尽管cELISA的特异性略低于iELISA,在犬布鲁氏菌病检测中仍然保持较高的准确性。在氨基酸序列QPIYVYPDDKNNLKEPTITGY中鉴定了mAbE10的表位，提示其作为布鲁氏菌病诊断抗原的潜力。总之,基于E10的cELISA为检测动物布鲁氏菌病提供了一种有效的手段，对于中国的DIVA诊断尤其重要，其中BP26突变疫苗被广泛使用。
