Axonal degeneration in the anterior insular cortex is associated with Alzheimer’s co-pathology in Parkinson’s disease and dementia with Lewy bodies - Translational Neurodegeneration

Modified Bielschowsky silver staining [] was used for the evaluation of axonal degeneration in the anterior insula. Prior to staining, all glassware was thoroughly rinsed with purified water for several hours or overnight and metals were avoided. All incubations were performed at cold temperature [

]. Free-floating 60-μm tissue sections were rinsed in distilled water followed by Tris-buffered saline and incubated in 20% silver nitrate solution for 30 min at 5 °C in the dark. Subsequently, non-evaporated ammonium hydroxide 30%–33% solution was slowly added to the sections until the color turned black, and then incubated for 30 min at 5 °C in the dark . Sections were then rinsed with evaporated ammonia-water for 10 min .

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