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. 2010 Mar 1;30(9):2206–2219. doi: 10.1128/MCB.01181-09

FIG. 4.

FIG. 4.

Accumulation of GAKIN/KIF13B at axon tips. (A) Representative GAKIN/KIF13B-stained images of stage 1, stage 2, and stage 3 hippocampal neurons from E18 rats are indicated. The accumulation of GAKIN/KIF13B at the tips of neurites is indicated by the small white arrows. Bars, 20 μm. (B) Percentage of hippocampal neurons that have a single neurite (gray) or two or more neurites (black) containing GAKIN/KIF13B accumulation at stage 2 (n = 100) or stage 3 (n = 96). GAKIN/KIF13B accumulation is defined here as a level greater than 4 times the shaft intensity level. (C) Neurons at 3 DIV were stained for GAKIN/KIF13B and Tau-1 or MAP2. Merged images are also presented (GAKIN/KIF13B shown in green; Tau-1 and MAP2 shown in red). Bar, 20 μm. (D) (Top) Neurons were transfected with GFP and then stained for GAKIN/KIF13B at 3 DIV (green, GFP; red, GAKIN/KIF13B). Bar, 20 μm. (Bottom) The fluorescence intensity in the axon tip, shaft, and cell body was measured and is shown in the graph plot using line scan imaging (purple, GFP; blue, GAKIN/KIF13B). (E) Anti-GAKIN/KIF13B antibody was preincubated with GST (control) or GST-CAP-Gly immobilized on beads for 2 h. Neurons at 3 DIV were stained with absorbed anti-GAKIN/KIF13B antibody or IgG (control) (green) together with anti-Tau-1 antibody (red). Green and red signals at the axon tips are also shown separately. Bar, 20 μm. (F) Neurons at 3 DIV were stained for β-tubulin and GAKIN/KIF13B, and the growth cone area is magnified and indicated. A merged image is also shown (GAKIN/KIF13B [red]; β-tubulin [green]). Bar, 5 μm.