Immunoperoxidase detection of neuronal antigens in full-thickness whole mount preparations of hollow organs and thick sections of central nervous tissue.

摘要: Immunofluorescently stained whole mounts have proved useful for defining the innervation of gut and large blood vessels. Nerves supplying other hollow organs are usually studied in sections, which provide much less information. Aiming to describe entire rat uterus, we developed a method immunoperoxidase staining full-thickness that allowed us visualize all immunoreactive axons. Uterine horns were dissected out, slit open, stretched, pinned flat fixed. Entire treated with methanol/peroxide, buffered Triton X-100 normal serum then incubated primary antibodies, biotinylated secondary antibodies avidin-horseradish peroxidase (HRP), each at least 3 days. Peroxidase reactions revealed immunoreactivity. Immunostained dehydrated, infiltrated epoxy resin, mounted on slides under Aclar coverslips polymerized. We bladders, gut, major pelvic ganglia thick sections perfused medulla oblongata similarly assess applicability method. Using this method, could map uterine provided by axons variety antigens. also tyrosine hydroxylase-immunoreactive layers bladder, throughout 300 μm medulla. These observations show reliably reveals complete central nervous tissue. The has several advantages. resin-embedded tissue does not degrade; immunostaining is non-fading permanent neurochemically defined features can be mapped scale without confocal microscopy.