Knapp E, da Câmara Machado A, Pühringer H, Wang Q, Hanzer V, Weiss H, Weiss B, Katinger H, Laimer da Câmara Machado M
Institute of Applied Microbiology, School of Food and Biotechnology, University of Agriculture, Vienna, Austria.
J Virol Methods. 1995 Oct;55(2):157-73. doi: 10.1016/0166-0934(95)00033-q.
Immuno-tissue printing protocols for the localization of apple chlorotic leaf spot virus (ACLSV), stem grooving virus (SGV) and plum pox virus (PPV) in shoots of Prunus and Malus in vitro have been established for routine diagnosis in a virus elimination program. Since these viruses belong to different virus genera, the protocols were adapted according to the properties of the virus under investigation. Accumulation of ACLSV was highest in the base of the stem and decreased towards the apex of the shoots. ACLSV was found in the epidermis, the cortex, in the vascular bundles, but seldom in the pith tissue of in vitro apple shoots. ACLSV immuno-tissue printing was as sensitive as ELISA and the intensity of color signals in immuno-tissue prints correlated with absorbance values by two-step ELISA. SGV could be detected by immuno-tissue prints at infectivity levels, where it reacted negative in ELISA. SGV accumulated in the vascular bundles, occurred locally in the parenchymatic tissue, was found in high amounts in young leaves near the meristem, but not within the meristem. PPV was detected in all tissue types of stem sections with an irregular pattern reflecting the in vivo situation causing problems with detection. Discrimination of poorly and heavily infected shoots was possible with the naked eye.
已建立免疫组织印迹法，用于在体外李属和苹果属的嫩枝中定位苹果褪绿叶斑病毒（ACLSV）、茎沟病毒（SGV）和李痘病毒（PPV），以便在病毒清除计划中进行常规诊断。由于这些病毒属于不同的病毒属，因此根据所研究病毒的特性对方法进行了调整。ACLSV在茎基部的积累量最高，向嫩枝顶端逐渐减少。在体外苹果嫩枝的表皮、皮层、维管束中发现了ACLSV，但在髓组织中很少发现。ACLSV免疫组织印迹法与酶联免疫吸附测定（ELISA）一样灵敏，免疫组织印迹中的颜色信号强度与两步ELISA的吸光度值相关。在ELISA检测呈阴性的感染水平下，通过免疫组织印迹可以检测到SGV。SGV在维管束中积累，在薄壁组织中局部出现，在分生组织附近的幼叶中大量存在，但在分生组织内未发现。在茎段的所有组织类型中都检测到了PPV，其不规则模式反映了体内情况，给检测带来了问题。用肉眼就可以区分感染程度轻和重的嫩枝。
