Microtiter spectrophotometric biofilm production assay analyzed with metrological methods and uncertainty evaluation

The microbial biofilm is a structure often developed by microorganisms when performing their harmful effects, both in the medical and industrial fields. Therefore, methods allowing identification and analysis of the biofilm play a fundamental role in determining the kind of intervention needed to avoid these effects. The microtiter spectrophotometric assay is recognized as the gold standard method to quantify a biofilm and to analyze the anti-biofilm activity of various substances. The aim of the present work is to validate this method through an uncertainty evaluation, covering eight different microbial species.The results show that the microtiter spectrophotometric assay is adequate to perform the biofilm analysis, with a good reproducibility and a reasonable uncertainty. However, the method requires a thorough knowledge of bio-dynamics concerning microbial species tested, in order to perform some protocol improvements catering in turn for better results.

► The uncertainty pertaining a method used to evaluate bacterial biofilm was analyzed.
► The classic protocol shows a rather large uncertainty.
► An improvement of the method was therefore devised.
► Yielding better results in terms of uncertainty.