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Fig. 7. Microinjection of anti-NRP/B antibody inhibited the NGF-induced neuronal differentiation of PC12 cells. (A) PC12 cells were microinjected with purified anti-mouse IgG (panels a,b and c) or with purified anti-NRP/B antibody (panels d,e and f). pEGFP-N2 vector was co-injected as a marker. Microinjected cells were grown in the absence of NGF (panels a and d) or in the presence of NGF (50 ng/ml) (panels b,c,e and f) for 48 hours and then analyzed for morphological changes. Cells (c) and (f), in the same field as in (b) and (e), respectively, were analyzed using a fluorescent microscope. (B) Microinjection of anti-Csk antibody (as a negative control) and pEGFP-N2 into PC12 cells in the presence of NGF (50 ng/ml) for 48 hours. Microinjected cells in the same field were analyzed under a UV microscope (a) or a fluorescent microscope (b). Arrows indicate microinjected cells. (C) Quantitative analysis of neurite outgrowth upon microinjection of NRP/B antibodies. The percentage of cells with neurites either untreated or following treatment with NGF in the presence of IgG control or NRP/B antibody was calculated in representative experiments repeated three times. Data are the means±s.d. Bars, 10 µm.
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