Sample Preparation and Stereological Methods for the Study of Glomerular Ultrastructure Using Electron Microscopy

  • Kathryn E. WhiteEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 2067)


In this chapter we describe conventional methods used for preparing renal tissue for transmission electron microscopy. We also describe a relatively new technique, serial block face scanning electron microscopy. Protocols are given for processing, sectioning, and imaging of tissue along with methods for obtaining quantitative data from the results.

Key words

Glomerulus Mesangium Podocyte Stereology Electron microscopy 


  1. 1.
    Basgen JM et al (1988) Measuring the volume density of the glomerular mesangium. Nephron 50(3):182–186CrossRefGoogle Scholar
  2. 2.
    Gundersen HJ, Jensen TB, Osterby R (1978) Distribution of membrane thickness determined by lineal analysis. J Microsc 113(1):27–43CrossRefGoogle Scholar
  3. 3.
    Gundersen HJ, Osterby R (1977) Glomerular size and structure in diabetes mellitus. II. Late abnormalities. Diabetologia 13(1):43–48CrossRefGoogle Scholar
  4. 4.
    Osterby R (1973) A quantitative electron microscopic study of mesangial regions in glomeruli from patients with short term juvenile diabetes mellitus. Lab Investig 29(1):99–110PubMedGoogle Scholar
  5. 5.
    Osterby R (1972) Morphometric studies of the peripheral glomerular basement membrane in early juvenile diabetes. I. Development of initial basement membrane thickening. Diabetologia 8(2):84–92CrossRefGoogle Scholar
  6. 6.
    White KE, Bilous RW (2000) Type 2 diabetic patients with nephropathy show structural-functional relationships that are similar to type 1 disease. J Am Soc Nephrol 11(9):1667–1673PubMedGoogle Scholar
  7. 7.
    White KE et al (2002) Podocyte number in normotensive type 1 diabetic patients with albuminuria. Diabetes 51(10):3083–3089CrossRefGoogle Scholar
  8. 8.
    Mauer M et al (2015) Glomerular structural-functional relationship models of diabetic nephropathy are robust in type 1 diabetic patients. Nephrol Dial Transplant 30(6):918–923CrossRefGoogle Scholar
  9. 9.
    (2001) Effect of 3 years of antihypertensive therapy on renal structure in type 1 diabetic patients with albuminuria: the European Study for the Prevention of Renal Disease in Type 1 Diabetes (ESPRIT). Diabetes 50(4):843–850Google Scholar
  10. 10.
    Gilbert RE, Cooper ME (1999) The tubulointerstitium in progressive diabetic kidney disease: more than an aftermath of glomerular injury? Kidney Int 56(5):1627–1637CrossRefGoogle Scholar
  11. 11.
    Steffes MW et al (2001) Glomerular cell number in normal subjects and in type 1 diabetic patients. Kidney Int 59(6):2104–2113CrossRefGoogle Scholar
  12. 12.
    Lemley KV et al (2000) Evolution of incipient nephropathy in type 2 diabetes mellitus. Kidney Int 58(3):1228–1237CrossRefGoogle Scholar
  13. 13.
    Maunsbach AB (1966) The influence of different fixatives and fixation methods on the ultrastructure of rat kidney proximal tubule cells. II. Effects of varying osmolality, ionic strength, buffer system and fixative concentration of glutaraldehyde solutions. J Ultrastruct Res 15(3):283–309CrossRefGoogle Scholar
  14. 14.
    Weibel ER, Bolender RP (1973) Stereological techniques for electron microscopy. In: Hayat MA (ed) Principles and techniques for electron microscopy. Van Nostrand Reinhold, New York, pp 273–296Google Scholar
  15. 15.
    Nyengaard JR (1999) Stereologic methods and their application in kidney research. J Am Soc Nephrol 10(5):1100–1123PubMedGoogle Scholar
  16. 16.
    White KE, Bilous RW (2004) Estimation of podocyte number: a comparison of methods. Kidney Int 66(2):663–667CrossRefGoogle Scholar
  17. 17.
    Weibel ER, Gomez DM (1962) A principle for counting tissue structures on random sections. J Appl Physiol 17:343–348CrossRefGoogle Scholar
  18. 18.
    Ichimura K et al (2015) Three-dimensional architecture of podocytes revealed by block-face scanning electron microscopy. Sci Rep 5:8993CrossRefGoogle Scholar
  19. 19.
    Randles MJ et al (2016) Three-dimensional electron microscopy reveals the evolution of glomerular barrier injury. Sci Rep 6:35068CrossRefGoogle Scholar
  20. 20.
    Lemley KV et al (2013) Estimation of glomerular podocyte number: a selection of valid methods. J Am Soc Nephrol 24(8):1193–1202CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2020

Authors and Affiliations

  1. 1.EM Research Services, Faculty of Medical SciencesNewcastle UniversityNewcastle upon TyneUK

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