Comparison of SEM and VPSEM imaging techniques with respect to Streptococcus mutans biofilm topography

Kathryn Weber, Juliana Delben, Timothy G. Bromage, Simone Duarte

Research output: Contribution to journalArticlepeer-review


The study compared images of mature Streptococcus mutans biofilms captured at increasing magnification to determine which microscopy method is most acceptable for imaging the biofilm topography and the extracellular polymeric substance (EPS). In vitro S. mutans biofilms were imaged using (1) scanning electron microscopy (SEM), which requires a dehydration process; (2) SEM and ruthenium red (SEM-RR), which has been shown to support the EPS of biofilms during the SEM dehydration; and (3) variable pressure scanning electron microscopy (VPSEM), which does not require the intensive dehydration process of SEM. The dehydration process and high chamber vacuum of both SEM techniques devastated the biofilm EPS, removed supporting structures, and caused cracking on the biofilm surface. The VPSEM offered the most comprehensive representation of the S. mutans biofilm morphology. VPSEM provides similar contrast and focus as the SEM, but the procedure is far less time-consuming, and the use of hazardous chemicals associated with SEM dehydration protocol is avoided with the VPSEM. The inaccurate representations of the biofilm EPS in SEM experimentation is a possible source of inaccurate data and impediments in the study of S. mutans biofilms.

Original languageEnglish (US)
Pages (from-to)175-179
Number of pages5
JournalFEMS Microbiology Letters
Issue number2
StatePublished - Jan 2014


  • Biofilm
  • Extracellular polymeric substance
  • SEM
  • Streptococcus mutans

ASJC Scopus subject areas

  • Microbiology
  • Molecular Biology
  • Genetics


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