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Scientifica
Volume 2014 (2014), Article ID 231083, 6 pages
http://dx.doi.org/10.1155/2014/231083
Research Article

Biofilm Localization in the Vertical Wall of Shaking 96-Well Plates

LEPABE, Department of Chemical Engineering, Faculty of Engineering, University of Porto, Rua Dr. Roberto Frias, 4200-465 Porto, Portugal

Received 20 November 2013; Accepted 19 December 2013; Published 13 April 2014

Academic Editors: M. Rodrigues and A. Sellam

Copyright © 2014 Luciana C. Gomes et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Microtiter plates with 96 wells are being increasingly used for biofilm studies due to their high throughput, low cost, easy handling, and easy application of several analytical methods to evaluate different biofilm parameters. These methods provide bulk information about the biofilm formed in each well but lack in detail, namely, regarding the spatial location of the biofilms. This location can be obtained by microscopy observation using optical and electron microscopes, but these techniques have lower throughput and higher cost and are subjected to equipment availability. This work describes a differential crystal violet (CV) staining method that enabled the determination of the spatial location of Escherichia coli biofilms formed in the vertical wall of shaking 96-well plates. It was shown that the biofilms were unevenly distributed on the wall with denser cell accumulation near the air-liquid interface. The results were corroborated by scanning electron microscopy and a correlation was found between biofilm accumulation and the wall shear strain rates determined by computational fluid dynamics. The developed method is quicker and less expensive and has a higher throughput than the existing methods available for spatial location of biofilms in microtiter plates.