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|Session Title: Aging and Stress Session Type: Poster|
|Session Location: Pauley Pavilion Session Time: Sunday - Tuesday|
|Poster Board Number: 409C Presentation Time: TUE, June 28, 2005, 3:00-4:30PM|
|Keywords: KW65 - Stress response|
Development of medium-throughput toxicity screens using C.
elegans. Windy A. Boyd, Sandra J. McBride, Jonathan H.
Freedman. Nicholas School of the Environment and Earth Sciences, Duke
Univ, Durham, NC.|
The National Toxicology Program (NTP) is responsible for the development of sound scientific tests designed to estimate the effects of chemicals on human health. Recently, the NTP and other regulatory agencies have recognized the need for alternative toxicological methods and models to decrease the time and expense of current toxicity testing protocols. The numerous advantages of using C. elegans as a model organism are well documented. Recently, these advantages have led to a rise in the use of C. elegans as a toxicity testing organism. Short life cycles, easy and inexpensive maintenance and culturing, and detailed biological knowledge allow for the development of rapid, low-cost toxicity tests that readily lend themselves to mechanistic studies of toxicant actions. Through collaboration with the NTP, our lab is now developing the means to screen approximately 200 potential neurological and developmental toxicants using C. elegans.
To this point, sublethal toxicity endpoints including growth, reproduction, movement, and feeding are automated using two liquid handling robotic workstations; a Complex Object Parametric Analyzer and Sorter (COPAS) BIOSORT for dispensing and analyzing worm length and fluorescence; and an imaging workstation for motion tracking and multidimensional image analysis. To optimize the number of chemicals screened, 96-well plate formats are used for sample preparation, dispensing of test organisms, and quantification of specific toxicological endpoints. As worms are dispensed, the time of flight, extinction, green, and red fluorescence are measured for each worm by the COPAS BIOSORT. Worms are exposed to toxicants over specific developmental stages for each test: L1s for 72-h growth, L4s for 48-h reproduction, and 3-day-old adults for 4-h or 24-h movement and feeding assays. After toxicant exposures for growth, reproduction, or feeding, the samples are aspirated with the COPAS REFLX and the same four parameters are measured. Movement tracking and image capturing of worms are also performed after toxicant exposures. For each test, the effective concentration that results in a 50% reduction in response relative to controls (EC50) is then calculated. Many chemicals have been tested and used as model test chemicals for development of the screens. Cadmium data will be presented as illustrations of the methods and results of each assay.
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