, Volume 97, Issue 2, pp 195–199 | Cite as

Effects of tissue processing techniques in acoustical (1.2 GHz) and light microscopy

  • A. F. W. van der Steen
  • J. M. Thijssen
  • G. P. J. Ebben
  • P. C. M. de Wilde


In this study the influence of various tissue processing and staining techniques on the acoustical properties of liver tissue was investigated. A qualitative study was performed using ultrasound attenuation as the imaged parameter of a combined optical/acoustical microscope with a 1.2 GHz transducer. Images were made of three sets of adjacent liver sections (6 μm in thickness) which were prepared in ten different ways: fixed by alcohol or formalin; stained by hematoxylineosin (HE), toluidine blue (TB) or non-stained; sectioned by a cryostat or by a paraffin microtome. It was concluded that the images obtained from cryostat sections were of much higher quality than those from paraffin sections. Images obtained from sections that were sectioned while embedded in paraffin displayed no detail at all. No consistent effect was noticed with respect to staining by HE or TB. Alcohol fixed sections gave more detailed images than formalin fixed sections. Formalin fixation in combination with cryostat sectioning yielded many cytoplasmic vacuoles.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. D'Astous FT, Foster FS (1986) Frequency dependence of ultrasound attenuation and backscatter in breast tissue. Ultrasound Med Biol 12:795–808Google Scholar
  2. Bamber JC, Nassiri DK (1985) Effect of gaseous inclusions on the frequency dependence of ultrasonic attenuation in liver. Ultrasound Med Biol 11:293–298Google Scholar
  3. Bamber JC, Frey MJ, Hill CR, Dunn F (1977) Ultrasonic attenuation and backscattering by mammalian organs as a function of time after excision. Ultrasound Med Biol 3:15–20Google Scholar
  4. Bamber JC, Hill CR, King JA, Dunn F (1979) Ultrasonic propagation through fixed and unfixed tissues. Ultrasound Med Biol 5:159–165Google Scholar
  5. Crosby BC, Mackay RA (1978) Some effects of time post-mortem on ultrasonic transmission through tissue under different modes of handling. IEEE Trans Biomed Eng 25:91–92Google Scholar
  6. Daft CMW, Briggs GAD (1989) The elastic microstructure of various tissues. J Acoust Soc Am 85:416–422Google Scholar
  7. Daft CMW, Weaver JMR, Briggs GAD (1986) Tissue characterization with microscopic resolution. IEEE Ultrasonics Symposium (IEEE Publ. No. 86CH2375-4), pp 945–948Google Scholar
  8. Daft CMW, Briggs GAD, O'Brien WD Jr (1989) Frequency dependence of tissue attenuation measured by acoustic microscopy. J Acoust Soc Am 85:2194–2201Google Scholar
  9. Foster FS, Strban M, Austin G (1984) The ultrasonic macroscope: initial studies of breast tissue. Ultrasonic Imaging 6:243–261Google Scholar
  10. Frizell LA, Carstensen EL, Davis JD (1979) Ultrasonic absorption in liver tissue. J Acoust Soc Am 65:1309–1312Google Scholar
  11. Hafsteinsson H, Rizvi SSH (1984) Acoustic microscopy — Principles and applications in the studies of biomaterial microstructure. Scanning Electr Microsc 3:1237–1247Google Scholar
  12. Hildebrand JA, Rugar D (1984) Measurement of cellular elastic properties by acoustic microscopy. J Microsc 134:245–260Google Scholar
  13. Jipson V, Quate CF (1978) Acoustic microscopy at optical wavelengths. Appl Phys Lett 32:789–791Google Scholar
  14. Kanngiesser H, Anliker M (1991) Ultrasound microscopy of biological structures with weak reflecting properties. In: Ermert H, Harjes HP (eds) Acoustical imaging, vol 19. Plenum, New York, London (in press)Google Scholar
  15. Kessler LW (1973) VHF ultrasonic attenuation in mammalian tissue. J Acoust Soc Am 53:1759–1760Google Scholar
  16. Kolodziejczyk E, Saurel JM, Bagnol J, Attal J, Fernandez-Graf MR, Saied A (1988) Transmission acoustic microscopy of tissue section (1 GHz). Histoacoustics and acoustic staining. Histochemistry 88:165–169Google Scholar
  17. Lemons RA, Quate CF (1974) Acoustic microscope-scanning version. Appl Phys Lett 24:163–165Google Scholar
  18. Litniewski J, Bereiter-Hahn J (1990) Measurements of cells in culture by scanning acoustic microscopy. J Microsc 185:95–107Google Scholar
  19. O'Brien WD (1977) The role of collagen in determining ultrasonic propagation properties in tissue. In: Kessler LW (ed) Acoustic holography, vol 7. Plenum, New York, pp 37–50Google Scholar
  20. O'Donnell M, Mimbs JW, Miller JG (1981) Relationship between collagen and ultrasonic backscatter in myocardial tissue. J Acoust Soc Am 69:580–588Google Scholar
  21. Okawai H, Tanaka M, Dunn F, Chubachi N, Honda K (1989) Quantitative display of acoustic properties of the biological tissue elements. In: Shimizu H, Chubachi N, Kushibiki J (eds) Acoustical imaging, vol 17. Plenum, New York London, pp 193–201Google Scholar
  22. Okawai H, Tanaka M, Dunn F (1990) Non-contact acoustic method for the simultaneous measurement of thickness and acoustic properties of biological tissues. Ultrasonics 28:401–410Google Scholar
  23. Parker KJ (1983) Ultrasonic attenuation and absorption in liver tissue. Ultrasound Med Biol 9:363–369Google Scholar
  24. Steen AFW van der, Cuypers MHM, Thijssen JM, Wilde PCM de (1991a) Influence of histochemical preparation on acoustic parameters of liver tissue, a 5 MHz study. Ultrasound Med Biol (in press)Google Scholar
  25. Steen AFW van der, Cuypers MHM, Thijssen JM, Ebben GPJ, Wilde PCM de (1991b) Preparation techniques in acoustical and optical microscopy of biological tissues, a study at 5 MHz and 1.2 GHz. In: Ermert H, Harjes HP (eds) Acoustical imaging, vol 19. Plenum, New York London (in press)Google Scholar

Copyright information

© Springer-Verlag 1992

Authors and Affiliations

  • A. F. W. van der Steen
    • 1
  • J. M. Thijssen
    • 1
  • G. P. J. Ebben
    • 2
  • P. C. M. de Wilde
    • 2
  1. 1.Biophysics Laboratory, Institute of OphthalmologyUniversity Hospital, St. RadboudNijmegenThe Netherlands
  2. 2.Department of PathologyUniversity Hospital, St. RadboudNijmegenThe Netherlands

Personalised recommendations