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Fig. 2. The distribution of A-GFP is similar to that of endogenous SCG10 in primary hippocampal neurons. (A) After 5 days in culture, primary hippocampal neurons were transfected with domain A of SCG10 fused to GFP (A-GFP). The fusion protein and the endogenous SCG10 were detected using monoclonal anti-GFP or polyclonal anti-SCG10 antibodies, respectively. Left panel is a low-magnification view of a transfected neuron showing the localization of A-GFP at a perinuclear compartment and along all processes up to their growth cone tips (asterisks). The three images to the right show a high magnification detail of a neuron double-labelled for A-GFP and endogenous SCG10, with the corresponding overlay image showing the colocalization of A-GFP and SCG10 in the perinuclear, Golgi-like region. All images are projections of confocal stacks. (B) Domains A of SCG10, SCLIP and RB3 were fused to GFP and transfected into HeLa cells. In all cases, GFP fluorescence showed a perinuclear as well as vesicular distribution, similar to the subcellular localization described for the corresponding stathmin-related proteins (Gavet et al., 1998). Bar, 10 µm.
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