A General-Purpose Silver Technique for Peripheral Nerve Fibers in Frozen Sections

Abstract

For the study of peripheral nerve fibers and endings, frozen sections of material fixed for 1 wk to 2 mo in Richardson's formalin, (prepared from paraformaldehyde and containing 3.4% sucrose) are extracted in saturated boric acid for 1–18 hr at 37° C, or in formol-alcohol (4 parts absolute ethanol, 1 part 20% formaldehyde) for 1–2 hr at 37° C. Tissues fixed for 3 mo or more do not require extraction. The sections are placed in 10% AgNo₃ for 20–25 min at 37° C, then passed through 4 baths of 10% formaldehyde during 15–30 min. They are developed in a diammino-silver solution prepared from Ag₂CO₃, or from AgNO₃, fixed for 10–30 min in 5% Na₂SO₃, containing 5% ammonia, washed well, toned for 10 min in 0.5% gold chloride, washed briefly and fixed in 5% Na₂S₂O₃. Finally they are washed, dehydrated, cleared, and mounted in Canada balsam.