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目的:检测和比较6种针对大鼠Nogo-A分子不同表位的单克隆抗体(mAb)应用于中枢神经组织的免疫组化染色特性,从而决定它们今后可能的应用范围和方法。其中4种mAb是针对Nogo-A分子中Nogo66片段,分别命名为No-go66-1、Nogo66-2、Nogo66-3和Nogo66-4;其他2种mAb是针对Nogo-A分子氨基端(570-691)肽段,分别为Nogo-N1和Nogo-N2。方法:利用免疫荧光组织化学,检测Nogo-A mAb抗体对大鼠神经组织的反应性与特异性。结果:正常SD大鼠脊髓组织的免疫荧光组织化学双标结果显示,制备的6种抗大鼠Nogo-A分子mAb均可以与商品化的兔抗大鼠Nogo-A多克隆抗体(PcAb)双标记;其中Nogo66-1、Nogo66-2、No-go66-4和NogoN-2 mAb可与MBP阳性细胞双标记,与GFAP阳性细胞不共存;而Nogo66-3和NogoN-1 mAb不仅可以与MBP阳性细胞双标,同时也可以和GFAP阳性细胞共存。结论:Nogo66-1、Nogo66-2、Nogo66-4和NogoN-2 mAb可很好的识别组织中的Nogo-A分子,用于免疫组化研究。
OBJECTIVE: To detect and compare the immunohistochemical staining of six kinds of monoclonal antibodies (mAbs) targeting different epitopes of rat Nogo-A in central nervous system to decide their possible future application scope and methods. Four of these mAbs were directed against the Nogo66 fragment of Nogo-A, named No-go66-1, Nogo66-2, Nogo66-3 and Nogo66-4, respectively; the other two mAbs were directed against the N-terminal of Nogo- 691) peptides, Nogo-N1 and Nogo-N2, respectively. Methods: Immunofluorescence histochemistry was used to detect the reactivity and specificity of Nogo-A mAb in rat neural tissue. Results: Immunofluorescence histochemical double-labeled results of spinal cord of normal SD rats showed that the prepared 6 kinds of anti-rat Nogo-A molecule mAb can be used in combination with commercial rabbit anti-rat Nogo-A polyclonal antibody (PcAb) The Nogo66-1, Nogo66-2, No-go66-4 and NogoN-2 mAb were double-labeled with MBP-positive cells and did not coexist with GFAP-positive cells. Nogo66-3 and NogoN-1 mAbs could not only be positive with MBP Double-labeled cells can also co-exist with GFAP-positive cells. Conclusion: Nogo66-1, Nogo66-2, Nogo66-4 and NogoN-2 mAb can well identify Nogo-A molecules in tissues for immunohistochemistry.