The observation of peroxisomes in NIE-115 neuroblastoma cells.

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Cultured neuroblastoma cells were transfected with a plasmid for GFP with a peroxisome targeting signal, SKL, in its C-terminal in order to visualize and characterize peroxisome in nerve cells. Materials and Methods: A plasmid containing a cDNA for GFP with SKL tripeptide in its C-terminal with CMV promoter (pEGFP-SKL) was constructed. NIE-115 neuroblastoma cells were cultured in DMEM medium supplemented with 10% FBS in 5% CO_2. They were transfected with pEGFP-SKL by calcium phosphate method. The medium was changed to that without FBS in order to observe neural processes. They were observed by an inverted fluorescent microscope equipped with a digital cooled CCD camera. After fixation in 4% peraformaldehyde, they were stained by immunofluorescent method with antibody 'for catalase, a marker enzyme for peroxisorne. Results: Fluorescent granules, 0.1 to 0.3 μm in their diameter, were observed in the cell body and neural processes under the serum starvation condition. As they were immunopositive for catalase, they were confirmed to be peroxisomes. Movement was not observed in most peroxisome, small number of them showed slow or fast movement in neural processes. Conclusion: The localization and movement of peroxisomes in cultured nerve cells were made possible by employing a method of transfection with green fluorescent protein.
日本組織細胞化学会の論文