IN VIVO LABELING OF MAMMALIAN CONE PHOTORECEPTORS BY INTRAVITREAL INJECTION OF FLUORESCENTLY TAGGED PEANUT AGGLUTININ

摘要

A method for specifically labeling outer and inner segments and pedicles of mammalian cone photoreceptors in vivo comprising Intravitreal injection of 2 μl for adult anesthestized mouse (C57BL/6 J), 10 μl for guinea pig (Duncan Hartley) of 20 μl for monkey (Macaca mulatta) of PNA conjugated with a fluorescent probe (Rhodamine or Fluorescein) at a concentration of 0.005%, 0.01%, 0.02%, 0.05% or 0.5% (only 0.5% for guinea pig and monkey) with a glass electrode with tip diameter of ~ 1 μm or an insulin syringe posterior to the limbus into the vitreous of the eye, Removing the eye under anesthesia at various time points (mouse: 0.5, 1, 2, 4 and 8 hours; 1, 2, 4, 7 and 10 days; 1 and 3 months; guinea pig and monkey: 30 minutes) after the PNA injection, Hemisecting and fixing the posterior eye-cup in 4% paraformaldehyde for 1 hour at 4°C, Flat mounting the retina on a glass slide or cryosectioning, and observing PNA labeling with a fluorescence microscope.