Question

Give a detailed description of the Ames assay including how it is performed, the basic principle of how mutants are detected, what general types of mutations can be detected, and the advantages and disadvantages of the Ames assay over more complex mammalian genotoxicity tests.

Answer 1

- The Ames assay test is used to check whether a given chemical can cause any mutation to the DNA of the test organism.

- It uses several strains of the bacterium Salmonella typhimurium that carry mutations in genes involved in histidine synthesis

- The bacteria are spread on an agar plate with a small amount of histidine.

- This small amount of histidine in the growth medium allows the bacteria to grow for initial time and have the opportunity to mutate.

- When the histidine is depleted only bacteria that have mutated to gain the ability to produce its own histidine will survive.

- The plate is incubated for 48 hours.

- The mutagenicity of a substance is proportional to the number of colonies observed.

- The ames test can find out whether the mutation is frameshift or point mutation

Advantages:

The advantage of this technique is that it serves as a quick and convenient assay to estimate the carcinogenic potential of a compound because standard carcinogen assays on mice and rats are time-consuming (taking two to three years to complete) and expensive

Disadvantages:

- Test can report false-positives and false-negatives

- Secondly carcinogens act by reacting with genetic material. The type which are metabolically activated generally form free radicals which then react with DNA. Many carcinogens have an affinity for a specific sequence of nucleotides. Obviously the number of sequences that are shared between mammals and bacteria are going to be very small! There may well be many chemicals which cause mutations in bacteria but which have absolutely no effect on mammals.

- carcinogens are divided into those which require to be metabolized in a cell (activated) and those which don't. Obviously the metabolism of a bacterial cell is going to differ from that of a mammalian cell. Compounds which are activated in a bacterial cell may therefore show no activity in a mammalian cell and vice versa