Beyond mortar and pestle grinding: A simple method to process bioanalytes collected by water filtration
Paparini, A. (2012) Beyond mortar and pestle grinding: A simple method to process bioanalytes collected by water filtration. In: Australian Society for Microbiology 2012 Annual Scientific Meeting, 1 - 4 July 2012, Brisbane, QLD.
The study of aquatic ecosystems, often involves the analysis of (bio)chemical compounds isolated from planktic microorganisms and particulate. These analyses require: concentration by filtration, optional detachment of the material from the support, disruption of cells and extracellular matrix, followed by chemical extraction, and quantification. Mortar/pestle are commonly used to powder the filter and its content, bypassing detachment. Grinding has low through-put capacity, and is time-consuming, cumbersome, and prone to cross-contaminations, when the same mortar/pestle are used for multiple isolations.
Objective. To develop a simple, rapid, and cheap procedure circumventing the need for detachment and disruption by mortar/pestle, while proving reliable and valid, in comparison to other methods.
Method. Lacustrine and estuarine water was filtered to concentrate seston material. Liquid nitrogen and glass beads in sterile single-use tubes, were used for filter powdering by a novel manual shaking method. Three alternative strategies (mortar and pestle, filter washing, automatic bead-beating) were implemented for comparison. Total proteins, extracted by organic solvents and quantified colorimetrically, were used as indicator of efficiency.
Results. The novel manual shaking method was the fastest and most through-put procedure. It consistently provided the highest yield (p<0.002 in pond water), costing only ~US$ 0.55/sample, for materials. The least effective methods, for pond– and estuarine–water respectively, were automatic bead-beating (42.2% vs. manual shaking), and filter washing (73.4%).
Conclusions. Without mortar/pestle grinding, this simple procedure provided comparable–, or significantly–higher yields than other methods. With minor modifications of the filtration and extraction steps, the method is suitable to analyse additional compounds.
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|Murdoch Affiliation:||School of Veterinary and Biomedical Sciences|
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