Figure 1
Adaptation of Golgi and Golgi-Cox staining for high-resolution LM and EM analysis. Pink, workflow for Golgi-Cox staining for light microscopy. Mice are perfused with PFA at day 1. After postfixation for 1 h in the same fixative, vibratome sections (100–500 µm) of the brain are made. Thin section (100 µm) could be impregnated with a sufficient number of neurons after 15 days in Golgi-Cox solution. Thick sections (150–500 µm) require 22–36 days, and whole brain or brain blocks, 40–45 days. The ammonium hydroxide step is made at day 15 for thin sections, day 23–36 for thick sections, or day 40–45 for the brain or brain blocks. Blue, workflow for original Golgi staining for light and electron microscopy. Mice are perfused as in above, and samples are postfixed for 24 h, followed by 10 days of impregnation with potassium dichromate. From day 12, samples are submitted to silver nitrate solution for at least 3 days, followed by gold toning for 40–60 minutes. Sections can be imaged by light microscopy from 15 days on. The EM preparation and embedding procedures take another 5–6 days. Purple, workflow for tissue clearing upon Golgi-Cox and original Golgi staining. For brain or large brain blocks after Golgi-Cox staining, the duration of clearing is 8 days, for hippocampi overnight, for thick (300–500 µm) sections between 2 h and overnight, and for thin sections 10 min. Original Golgi-stained sections and brain blocks required longer clearing procedures, 1 and 21 days respectively. See text for details. Abbreviations: EM, electron microscopy; LM, light microscopy; PFA, paraformaldehyde; PB, phosphate buffer; TEM, transmission electron microscopy.
