Purification and Characterization of Neurofilament

純化及鑑定神經纖維絲之研究

洪秀貞(Show-Jen Hong) ； 柯立文(Li-Wen Ko) ； 陳英俊(Ing-Jun Chen)

《The Kaohsiung Journal of Medical Sciences》 4卷2期 (1988/02) Pp. 63-71

https://doi.org/10.6452/KJMS.198802.0063

摘要

本研究是利用免疫化學法觀察神經系統特有之中間絲（imtermediate filament）神經纖維絲（neurofilament）在坐骨神經，脊髓，視神經及視交叉內的分怖形態。首先利用牛腦白質經過 hydroxylapatite 層析分離法及SDS膠質電泳法分離並純化神經纖維絲之成分蛋白質，接著以免疫實驗兔製備免疫血清，而以CNBr-Sepharose 4B 親和性層析法製備免疫親和性單價血清。本實驗為了證明免疫血清的特異性，我們利用免疫酵素法檢視經神經生長因子（nerve growth factor）處理後的嗜鉻性細胞瘤（PC12細胞）時發現：神經纖維絲成束聚集的出現在細胞質內。因此證明此神經纖維絲之抗體已製備成功，同時又利用此免疫血清配合免疫化學法觀察不同神經組織內神經纖維絲之分怖及神經纖維的構造形態。

並列摘要

The distribution of neurofilament was investigated in the sciatic nerve, spinal cord, optic nerve and optic chiasm by the immunocytochemical methods. The neurofilament protein (150 KDa) was purified from the white matter of bovine brains by the hydroxylapatite chromatography, followed by the SDS-polyacrylamide gel electrophoresis. Protein of neurofilament was eluted from the SDS-polyacrylamide gel, and the antibody was raised in rabbits. Immune serum was subjected to affinity chromatography on its corresponding antigen covalent-coupled to CNBr-Sepharose 4B. The specificity of neurofilament was proved by the immunoperoxidase staining in the nerve growth factor (NGF)-treated PC12 cells. Subsequently immunocytochemical examination of nervous system revealed that the antiserum of neurofilament bound to neuronal axons of sciatic nerve, spinal cord, optic nerve and optic chiasm. These results revealed that the monovalent antibody against neurofilament was prepared in success in our laboratory. And the topographical distribution of neuronal pathways were identified by the immunofluorescent and immunoperoxidase methods.

並列關鍵字

neurofilament ； intermediate filament ； nerve growth factor