KENBIKYO Vol.49▶No.2 2014
■Researches Today

Rapid Three-Dimensional Analysis of Renal Biopsy Paraffin Sections by Low Vacuum SEM

Sumire Inaga, Toshio Kameie, Hironobu Nakane, Tomonori Naguro and Toshiyuki Kaidoh

Abstract: Electron microscopic diagnosis of renal biopsy is conventionally performed by observation of ultra-thin sections of the renal tissue using a transmission electron microscope (TEM). However, it takes weeks to months to get a definitive diagnosis because of the long processing time. Furthermore, disadvantages of the use of TEM include sample size limitations and difficulty in obtaining three-dimensional information. Therefore, we developed a rapid three-dimensional analysis method using renal biopsy paraffin sections by low vacuum scanning electron microscopy (LVSEM) in the backscattered electron (BSE) mode. Unstained paraffin sections were deparaffinized, stained with platinum blue (Pt-blue) or periodic acid silver-methenamine (PAM), and directly observed with a table-top type LVSEM. Cut side views and surface views of glomeruli were shown under LVSEM. Pathological features could be investigated in some cases of human glomerular disease (minor glomerular abnormalities, IgA nephropathy, and membranous glomerulonephritis) at magnifications between ×50 to ×30,000. On Pt-blue-stained sections, podocyte, endothelium, mesangium and glomerular basement membrane (GBM) were clearly distinguished due to the different yields of BSE signals. PAM-positive GBM distinctly showed morphological alterations through overlying cells. LVSEM is expected to provide a novel approach to the pathological diagnosis of human glomerular diseases using conventional renal biopsy paraffin sections.

Key words: low vacuum SEM, renal biopsy paraffin sections, platinum blue (Pt-blue) staining, periodic acid silver-methenamine (PAM) staining, electron microscopic diagnosis