Russi Romina Cecilia, Garcia Lucila, Cámara María Silvia, Soutullo Adriana Rosa
Laboratorio de Diagnóstico e Investigaciones Agropecuarias., Ministerio de la Producción, Ciencia y Tecnología de la Provincia de Santa Fe, Santa Fe, Argentina.
Laboratorio de Inmunología Experimental, Cátedra de Inmunología Básica, Facultad de Bioquímica y Ciencias Biológicas, Universidad Nacional del Litoral, Santa Fe, Argentina.
Equine Vet J. 2023 Jan;55(1):111-121. doi: 10.1111/evj.13555. Epub 2022 Mar 1.
Equine infectious anaemia (EIA) is controlled by the identification of seropositive animals. The official diagnostic method is the agar gel immunodiffusion (AGID) test, which detects antibodies against a viral core protein (p26). Although AGID is inexpensive and specific, the report of results takes considerable time and the test has low analytical sensitivity.
To validate our in-house indirect ELISA , following the World Organization of Animal Health (OIE) criteria.
Test validation.
Synthetic peptides gp90 and gp45 were used as antigens in ELISA . Tests used for validation, calibration and linear working operating range, analytical and diagnostic sensitivity and specificity, repeatability and reproducibility were assessed by comparing them with the AGID test and using 1844 equine sera grouped into five different panels.
We were able to replace the National References Sera with our Internal Reference Sera. ELISA had acceptable repeatability and reproducibility. Analytical sensitivity of the ELISA was 800 times greater than that of AGID test for positive sera and 400 times greater for weak positive sera. ELISA also showed optimal analytical specificity, since no cross-reactivity was detected with antibodies against other equine viruses. One sample was positive by AGID test and negative by ELISAgp90/45. ELISA was performed using 243 EIA positive and 878 negative equid sera, and showed a diagnostic sensitivity of 99.59% [CI 97.73%-99.99%] and a diagnostic specificity of 90.32% [CI 88.17%-92.19%], compared to AGID test; thus, it was demonstrated to be a robust test.
Samples were derived from naturally infected equid populations showing heterogeneous clinical states: therefore, their status was uncertain and some horses were sampled more than once. The AGID test may not be the most useful gold standard.
ELISA is a useful tool for the diagnosis of EIAV infection and meets validation requirements established by the OIE.
马传染性贫血（EIA）通过识别血清学阳性动物来控制。官方诊断方法是琼脂凝胶免疫扩散（AGID）试验，该试验检测针对病毒核心蛋白（p26）的抗体。尽管AGID价格低廉且具有特异性，但结果报告需要相当长的时间，并且该试验的分析灵敏度较低。
按照世界动物卫生组织（OIE）的标准验证我们的内部间接ELISA。
试验验证。
合成肽gp90和gp45用作ELISA中的抗原。通过将其与AGID试验进行比较，并使用分为五个不同组的1844份马血清来评估用于验证、校准和线性工作范围、分析和诊断灵敏度及特异性、重复性和再现性的试验。
我们能够用内部参考血清替代国家参考血清。ELISA具有可接受的重复性和再现性。ELISA对阳性血清的分析灵敏度比AGID试验高800倍，对弱阳性血清高400倍。ELISA还显示出最佳的分析特异性，因为未检测到与针对其他马病毒的抗体的交叉反应。一个样本AGID试验呈阳性而ELISA gp90/45呈阴性。使用243份EIA阳性和878份阴性马血清进行ELISA，与AGID试验相比，诊断灵敏度为99.59%[CI 97.73%-99.99%]，诊断特异性为90.32%[CI 88.17%-92.19%]；因此，它被证明是一种可靠的试验。
样本来自表现出不同临床状态的自然感染马群：因此，它们的状态不确定，一些马被多次采样。AGID试验可能不是最有用的金标准。
ELISA是诊断马传染性贫血病毒感染的有用工具，符合OIE制定的验证要求。
