Rocke T E, Smith S R, Nashold S W
U.S. National Biological Survey, National Wildlife Health Center, Madison, Wisconsin 53711, USA.
J Wildl Dis. 1998 Oct;34(4):744-51. doi: 10.7589/0090-3558-34.4.744.
An enzyme-linked immunosorbent assay (ELISA) was developed for the detection of type C botulinum toxin (Clostridium botulinum) in wild birds. This simple, antigen-capture ELISA utilizes polystyrene immunosticks as the solid substrate, chicken antitoxin (IgY) as the coating antibody, rabbit antitoxin as the primary antibody, and peroxidase-labeled goat-anti-rabbit as the secondary antibody. To evaluate the immunostick ELISA as a diagnostic test for avian botulism, known concentrations of toxin were added to heparinized blood collected from healthy birds and tested by both the ELISA and mouse bioassay. Also, blood samples from 236 bird carcasses submitted to the National Wildlife Health Center (NWHC) for cause of death determinations were tested by both procedures. Using < or = 0.5 ml as the test volume for both procedures, the ELISA was less sensitive, detecting 0.25 ng/ml of toxin compared to 0.12 ng/ml for the mouse bioassay. Using the same volume of test sample for diagnostic submissions (< or = 0.5 ml), the ELISA was positive for 60% of the 149 clinically-diagnosed cases of botulism, whereas the mouse bioassay was positive for 79%. However, we demonstrated that with larger sample volumes (> or = 1.0 ml), the sensitivity of the ELISA may be equivalent or better than the mouse test due to the concentrating effect of the ELISA procedure. These preliminary results suggest that when adequate sample volumes are available, the immunostick ELISA can replace the mouse test for the diagnosis of botulism in wild birds.
已开发出一种酶联免疫吸附测定法（ELISA），用于检测野生鸟类中的C型肉毒杆菌毒素（肉毒梭菌）。这种简单的抗原捕获ELISA使用聚苯乙烯免疫棒作为固相载体，鸡抗毒素（IgY）作为包被抗体，兔抗毒素作为一抗，过氧化物酶标记的山羊抗兔抗体作为二抗。为了评估免疫棒ELISA作为禽肉毒中毒诊断试验的效果，将已知浓度的毒素添加到从健康鸟类采集的肝素化血液中，并通过ELISA和小鼠生物测定法进行检测。此外，还对提交给国家野生动物健康中心（NWHC）进行死因判定的236只鸟类尸体的血液样本进行了这两种检测。对于这两种检测方法，均使用≤0.5 ml作为检测体积，ELISA的灵敏度较低，检测到毒素的浓度为0.25 ng/ml，而小鼠生物测定法为0.12 ng/ml。对于诊断送检样本使用相同体积的检测样品（≤0.5 ml），ELISA对149例临床诊断为肉毒中毒的病例中的60%呈阳性，而小鼠生物测定法为79%。然而，我们证明，由于ELISA程序的浓缩作用，当使用更大的样本体积（≥1.0 ml）时，ELISA的灵敏度可能与小鼠试验相当或更好。这些初步结果表明，当有足够的样本体积时，免疫棒ELISA可替代小鼠试验用于诊断野生鸟类的肉毒中毒。
