IGEM:IMPERIAL/2007/Projects/Biofilm Detector/TestingValidation: Difference between revisions
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It is necessary to test the Infector Detector system in an environment similar to that of its intended operation, if not in the environment itself. It was found that biofilms can be modelled by a gel with pore size of 80nm and fiber thickness of 0.6nm<cite>1</cite>. Agar gel, prepared a concentration of 2% by weight has approximately these properties<cite>2</cite>. | It is necessary to test the Infector Detector system in an environment similar to that of its intended operation, if not in the environment itself. It was found that biofilms can be modelled by a gel with pore size of 80nm and fiber thickness of 0.6nm<cite>1</cite>. Agar gel, prepared a concentration of 2% by weight has approximately these properties<cite>2</cite>. | ||
Therefore, an agar gel was prepared at a concentration of 2% (by weight) and with AHL dissolved at different concentrations (3nM, 5nM, 7nM, 10nM). 40μl of these gel samples were then pipetted into wells in a 96-well fluorometer plate, according to the diagram on the left. Three of the 10nM AHL agar containing wells were further treated with 10μl of 10x diluted stock GFP solution. Additionally, several control wells were set up, containing: agar without AHL; agar without AHL and with GFP; GFP only; and | Therefore, an agar gel was prepared at a concentration of 2% (by weight) and with AHL dissolved at different concentrations (3nM, 5nM, 7nM, 10nM). 40μl of these gel samples were then pipetted into wells in a 96-well fluorometer plate, according to the diagram on the left. Three of the 10nM AHL agar containing wells were further treated with 10μl of 10x diluted stock GFP solution. Additionally, several control wells were set up, containing: agar without AHL; agar without AHL and with GFP; GFP only; and blanks (empty wells). | ||
Molten agar has a high surface tension, and is difficult to pipette and spread at the bottom of the plastic wells. Therefore, at least two wells were loaded with each sample, in order to determine the variability of measurements as a result of the well-loading procedure. | |||
Similarly, three wells were loaded with 10μl of GFP solution, because the small volume of the solution makes it difficult to spread the solution evenly at the bottom of the plate. | |||
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Revision as of 13:33, 18 September 2007
Infector Detector: Testing/Validation
Testing on Simulated Biofilms (Agar with AHL)
It is necessary to test the Infector Detector system in an environment similar to that of its intended operation, if not in the environment itself. It was found that biofilms can be modelled by a gel with pore size of 80nm and fiber thickness of 0.6nm[1]. Agar gel, prepared a concentration of 2% by weight has approximately these properties[2].
Therefore, an agar gel was prepared at a concentration of 2% (by weight) and with AHL dissolved at different concentrations (3nM, 5nM, 7nM, 10nM). 40μl of these gel samples were then pipetted into wells in a 96-well fluorometer plate, according to the diagram on the left. Three of the 10nM AHL agar containing wells were further treated with 10μl of 10x diluted stock GFP solution. Additionally, several control wells were set up, containing: agar without AHL; agar without AHL and with GFP; GFP only; and blanks (empty wells).
Molten agar has a high surface tension, and is difficult to pipette and spread at the bottom of the plastic wells. Therefore, at least two wells were loaded with each sample, in order to determine the variability of measurements as a result of the well-loading procedure.
Similarly, three wells were loaded with 10μl of GFP solution, because the small volume of the solution makes it difficult to spread the solution evenly at the bottom of the plate.
References
