Combined Staining Techniques for Demonstration of Staphylococcus aureus Biofilm in Routine Histopathology

Research output: Contribution to journalJournal articleResearchpeer-review

Aim: Visualization of Staphylococcus aureus biofilm using histochemical staining and combined histochemistry (HC) and immunohistochemistry (IHC).

Methods: The ability of S. aureus S54F9 to form biofilm was tested in vitro. Hereafter, infected bone tissue was collected from two different porcine models of osteomyelitis inoculated with S. aureus strain S54F9. The infection time was five and fifteen days, respectively. Twenty-five different histochemical staining protocols were tested in order to find the stains that could identify extracellular biofilm matrix. Protocols with an optimal visualization of biofilm extracellular matrix were combined with an immunohistochemical protocol based on a specific antibody against S. aureus. The combined protocols were applied to the tissue from the porcine models and to infected bone tissue from a child suffering from chronic staphylococcal osteomyelitis for more than a year.

Results: S. aureus S54F9 showed an ability to form biofilm in vitro. Visualization of biofilm, i.e. bacterial cells and extracellular matrix in different colours, was seen when the immunohistochemical protocol was combined with Alcian Blue pH3, Luna and Methyl-pyronin green. The bacterial cells were red to light brown and the extracellular matrix either light blue, blue or orange depending on the histochemical stain. In the porcine models and the human case 10 and 90 percent, respectively, of the bacterial aggregates in a 100x magnification field displayed both the extracellular matrix and the bacterial cells simultaneously in two different colours.

Conclusions: A combination of HC and IHC can be used to diagnose and characterise biofilm infections on a routine basis.
Original languageEnglish
JournalJournal of Bone and Joint Infection
Issue number1
Pages (from-to)27-36
Publication statusPublished - 2018

Number of downloads are based on statistics from Google Scholar and

No data available

ID: 192741486