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Experimental Protocols


Sample Size Considerations

In sampling a population of animals for this assay, it is useful to look at both juveniles and adults. The juvenile samples can indicate an immediate problem, whereas the adult samples can indicate a problem at some point in the animal's life history, the timing of which may not be obvious. In addition, the chemical signature of the mutagen may no longer be present in the animal.

It is useful, but not necessary to identify the sex of the samples. The body burden of man-made chemicals is unequal in the two sexes of animals that reproduce more than once. Females, through the production of eggs and young, can pass on much of their lipophilic chemical burden to their offspring, but the chemical burden of males keeps building throughout their lifespan.

The sample size of maximum sensitivity is 30 animals per sex per species per site for the mutagenicity assay. Where sexing is not possible (i.e. in juveniles or in live sampling of adults not in breeding season) then 30 animals per species per site is acceptable. Where the samples are going to be used as baseline data for future comparisons, then the samples need to be split into about 10 sub-samples. This methodology is described in the Protocol section below.

Sample Storage

For long term storage, all samples need to be kept in liquid nitrogen or in a -80oC freezer. Samples cannot be thawed out and then refrozen at any time. If this occurs then the samples will no longer be suitable for assay by flow cytometry.

Blood Collection Protocols

Fish less than 20 gm in weight

Fish greater than 20 gm in weight

Citrate Buffer - Freezing Media



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Updated by Michael Easton 2009.