Gaetano La Manna1, Gianandrea Pasquinelli2,3, Sabrina Valente4, Giorgia Comai1, Deborah Malvi5, Valeria Corradetti1. 1. Nephrology, Dialysis and Kidney Transplantation Unit, IRCCS, Azienda Ospedaliero-Universitaria di Bologna, Bologna, Italy. 2. Biotechnology and Methods in Laboratory Medicine, Department of Experimental, Diagnostic and Specialty Medicine (DIMES), University of Bologna, Bologna, Italy. 3. Subcellular Nephro-Vascular Diagnostic Program, Pathology Unit, IRCCS, Azienda Ospedaliero-Universitaria di Bologna, Bologna, Italy. 4. Biotechnology and Methods in Laboratory Medicine, Department of Experimental, Diagnostic and Specialty Medicine (DIMES), University of Bologna, Bologna, Italy. sabrina.valente2@unibo.it. 5. Pathology Unit, IRCCS, Azienda Ospedaliero-Universitaria di Bologna, Bologna, Italy.
Abstract
INTRODUCTION: Electron microscopy (EM) represents an indispensable technique for the diagnosis of kidney glomerular diseases. When dedicated tissue is not available, histological and cryostat sections can be reprocessed for EM using the pop-off technique. Here the practical value of this technique is analysed with emphasis on its accuracy in measuring basement membrane thickness and detecting immune deposits. METHODS: Ninety-four histological sections of kidney tissues fixed in Serra's solution, stained with H&E, PAS, and Masson's Trichrome; for EM analysis, the sections were recovered from either treated or untreated microscope slides through the pop-off technique. Some sections were recovered from cryosections allocated for immunofluorescence. RESULTS: The ultrastructural details were sufficiently maintained on tissues fixed with Serra's solution despite being considered disadvantageous for EM. The type of microscope slides and the time of biopsy storage did not affect the quality of section recovery. The histological stains had only moderate effects on the electron-density of the glomerular basement membrane (GBM). The pop-off technique reduced the GBM thickness when compared to the conventional EM processing but preserved the electron density of immune deposits. CONCLUSIONS: The application of the pop-off method to renal biopsy is a useful recovery method that produces limited but satisfactory results when there is no suitable material for EM. The ultrastructural morphology was retained even from tissues fixed with Serra's solution, and deposits maintained the expected electron density, however, we observed an overall thickness reduction of the GBM that could have a potential impact on thin membrane disease diagnosis.
INTRODUCTION: Electron microscopy (EM) represents an indispensable technique for the diagnosis of kidney glomerular diseases. When dedicated tissue is not available, histological and cryostat sections can be reprocessed for EM using the pop-off technique. Here the practical value of this technique is analysed with emphasis on its accuracy in measuring basement membrane thickness and detecting immune deposits. METHODS: Ninety-four histological sections of kidney tissues fixed in Serra's solution, stained with H&E, PAS, and Masson's Trichrome; for EM analysis, the sections were recovered from either treated or untreated microscope slides through the pop-off technique. Some sections were recovered from cryosections allocated for immunofluorescence. RESULTS: The ultrastructural details were sufficiently maintained on tissues fixed with Serra's solution despite being considered disadvantageous for EM. The type of microscope slides and the time of biopsy storage did not affect the quality of section recovery. The histological stains had only moderate effects on the electron-density of the glomerular basement membrane (GBM). The pop-off technique reduced the GBM thickness when compared to the conventional EM processing but preserved the electron density of immune deposits. CONCLUSIONS: The application of the pop-off method to renal biopsy is a useful recovery method that produces limited but satisfactory results when there is no suitable material for EM. The ultrastructural morphology was retained even from tissues fixed with Serra's solution, and deposits maintained the expected electron density, however, we observed an overall thickness reduction of the GBM that could have a potential impact on thin membrane disease diagnosis.
Authors: Giuseppe Cianciolo; Irene Capelli; Maria Cappuccilli; Roberto Schillaci; Mario Cozzolino; Gaetano La Manna Journal: Clin Kidney J Date: 2016-01-18