Nucleocapsid (N) protein is the most highly expressed of all avian metapneumovirus (aMPV) viral proteins and stimulates a substantial immune response in infected animals. Codon optimized recombinant N (rec-N) protein from aMPV subtypes A, B, and C were expressed using the baculoviral expression system in Trichoplusia ni (Tni) insect cells. A mixture of purified rec-N antigens from each subtype was used as a coating antigen and was evaluated in indirect ELISA (iELISA) to assess antibody response in serum samples collected from experimentally infected chickens and turkeys with different aMPV subtypes. Also, archived field serum samples that were collected from different poultry submissions were used. Receiver operating characteristic (ROC) analysis was performed using chicken and turkey serum samples that were confirmed by indirect fluorescent antibody (IFA) test for serostatus (positive n = 270, negative n = 610). The ROC analysis showed sensitivity and specificity of 97 % at a cut-off value of 0.25. The rec-N iELISA was compared with a commercial whole virus-based APV kit. The rec-N iELISA showed comparable results in detecting antibody response in aMPV infected chicken sera but was more sensitive in detecting early antibody response in aMPV infected turkey serum samples. Our results further confirm the presence of aMPV antibodies in Canadian domestic poultry populations. The developed aMPV-rec N iELISA offers a safe and valuable alternative to whole virus-based iELISA for serodiagnosis and seroepidemiological surveillance of the disease in domestic poultry.

核衣壳蛋白（N）是所有禽偏肺病毒（aMPV）病毒蛋白中表达最高的蛋白，可在感染的动物中刺激大量的免疫反应。使用杆状病毒表达系统在Trichoplusia ni中表达来自aMPV亚型A，B和C的密码子优化的重组N（rec-N）蛋白（Tni）昆虫细胞。来自每种亚型的纯化的rec-N抗原的混合物用作包被抗原，并通过间接ELISA（iELISA）进行评估，以评估从不同aMPV亚型的实验感染鸡和火鸡收集的血清样品中的抗体反应。此外，还使用了从不同家禽样品中收集的存档的现场血清样品。使用鸡和火鸡血清样品进行接收者操作特征（ROC）分析，这些样品通过间接荧光抗体（IFA）测试确定为血清状态（阳性n = 270，阴性n = 610）。ROC分析显示，临界值为0.25时，灵敏度和特异性为97％。将rec-N iELISA与市售的基于全病毒的APV试剂盒进行了比较。rec-N iELISA在检测被aMPV感染的鸡血清中检测抗体反应中显示出可比的结果，但在检测被aMPV感染的土耳其血清样品中早期抗体反应中更敏感。我们的结果进一步证实了加拿大家禽种群中存在aMPV抗体。所开发的aMPV-rec N iELISA提供了一种基于全病毒的iELISA的安全且有价值的替代方法，可用于家禽疾病的血清学诊断和血清流行病学监测。