Fig. 2: Histogram analysis to determine the optimal KMnO₄/Pb metal staining protocol.

Electron micrographs were randomly collected from the renal cortex and tilled for histogram analysis (Supplementary Fig. 3). Grayscale histograms depicting the number of pixels at each grey level, ranging from 0 to 255, present in each staining protocol. a All sections were preset for treatment with KMnO₄ for 5 min followed by Pb staining for 3 min. b All sections were preset for treatment with 0.2% KMnO₄ followed by Pb staining for 3 min. c No treatment; d 0.2% KMnO₄ for 5 min, alone; e 1% uranyl acetate for 5 min followed by Pb staining for 3 min; and f Pb staining for 3 min followed by 0.2% KMnO₄ for 5 min. Note the representative histogram after treatment with 0.2% KMnO₄ for 5 min followed by Pb staining for 3 min (a, b), corresponding to the conventional treatment with 1% uranyl acetate for 5 min followed by Pb staining for 3 min (e). N total number of pixels, Mean mean grey level, StdDev standard deviation.