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The silver staining protocol was taken from the publication of Novotny36. Briefly, the following steps were conducted:
PFA fixed, paraffin embedded sections are dewaxed and hydrated in distilled water via descending alcohols. Pretreated samples were incubated with 30% silver nitrate overnight, reduced in reducing solution, incubated in ammoniacal silver nitrate for 15 min., reduced second time and incubated for 5 min. in 5% sodium thiosulfate. After dehydrating via ascending alcohols, terpineol-xylene the samples were embedded in DePeX.
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