In vitro, test systems belong to four types: peripheral blood Ieukocytes, of man or other mammals, and primary or secondary cell lines isolated from biopsies of man and other mammals.
Criteria for evaluating the cytogenetic damage are: cell toxicity, chromosome breakage, including genome mutations and sister chromatid exchanges (SCE). The standardization of the tests involves rigorous definition of culture media and conditions, origin of cell lines, dose and purity of substances added to media: PHA, BUdR, colchicine and antibiotics, conditions of treatments (doses and purity of tested chemicals, manufacturers, solvents, etc.). The time following treatment at which damage is observed should be specified in relation to the peak of mitotic activity, due to mitotic inhibition increasing with dose.
All clastogenic effects should be recorded at metaphase but preferentially those with obvious genetical implications. The sensitivity of mammalian cells to well known mutagens is particularly high, especially for bifunctional alkylating agents. Threshold values and dose-response curves can be investigated and statistically evaluated. The predictability can be increased by correlating chromosome aberrations with SCE. There are examples however where discrepancies exist.
The stability of cell lines and the variations of the donors, especially lymphocytes could affect the reproducibility by modifying the control level of chromosome aberrations and the sensitivity. Modifications could also arise from an insufficient standardization and also the time of cell fixation after treatment and onset of cell culture.
The in vitro test systems can be limited by the occurrence of false negatives as in the case of cyclophosphamide and other mutagens carcinogens such as benzo-a-pyrene and nitrosamines, or false positives as in the case of some xanthine derivatives. These drugs pose the problem of evaluating the detoxication/activation capacity, for which a model has been worked out with several microsomal fractions.
In the sequence of tests, chromosome breakage (including SCE) observed in mammalian cells is obviously at a first step of mutagenicity