Determination Of Cell Aggregation In Bacteria Cultures With An Automatic And Reliable Method

By Andres Lobeiras, Beckman Coulter, Inc

The percentage of aggregates in bacteria cultures may be determined by employing the Coulter Principle, also known as Electrical Sensing Zone (ESZ) technique. The ISO International Standard 13319 describes this technique. A suitable electrolyte solution is required to perform the analysis, in many cases the Phosphate Buffered Saline (PBS) solution used to suspend the cells may be used as the electrolyte. The sample is prepared and then analyzed using a Beckman Coulter Multisizer 3 to determine the size distribution of the cells. The results are reported as number percent of aggregates.

The use of the Beckman Coulter™ Multisizer™ 3 provides a fast, easy, accurate and automatic method to determine cell aggregation. This instrument also provides reliable results not dependent of the operator’s judgment as opposed to results reported when using a hemocytometer.

Contact lens care disinfecting products must be evaluated for their safety and effectiveness. Procedures used to evaluate the efficacy of lens care disinfection solutions routinely employ a specific panel of potentially pathogenic challenge organism suspensions. Microscopic examination of challenge organism suspensions has demonstrated the presence of cell clumps. These aggregates prevent uniform exposure to the test disinfectant and result in underestimation of the true number of challenge organisms. In order to reduce the number of clumps, and therefore produce a more uniform cell suspension, some methods suggests filtration of bacterial challenge organism suspensions prior to use as test inoculate. The method also allows the use of suspension mediums containing 0.05% Tween 80 as a dispersant. The Multisizer 3 Coulter Counter may be used to conduct analysis on cell suspensions pre- and post-filtration to evaluate filtration effectiveness by determining the size distribution and relative number of cell clumps in the bacterial cell suspension. The effectiveness of the Tween 80 may also be evaluated.