Abstract
To comply with guiding principles for the ethical use of animals for experimental research, the field of mutation research has witnessed a shift of interest from large-scale in vivo animal experiments to small-sized in vitro studies. Mutation assays in cultured cells of transgenic rodents constitute, in many ways, viable alternatives to in vivo mutagenicity experiments in the corresponding animals. A variety of transgenic rodent cell culture models and mutation detection systems have been developed for mutagenicity testing of carcinogens. Of these, transgenic Big Blue® (Stratagene Corp., La Jolla, CA, USA, acquired by Agilent Technologies Inc., Santa Clara, CA, USA, BioReliance/Sigma-Aldrich Corp., Darmstadt, Germany) mouse embryonic fibroblasts and the λ Select cII Mutation Detection System have been used by many research groups to investigate the mutagenic effects of a wide range of chemical and/or physical carcinogens. Here, we review techniques and principles involved in preparation and culturing of Big Blue® mouse embryonic fibroblasts, treatment in vitro with chemical/physical agent(s) of interest, determination of the cII mutant frequency by the λ Select cII assay and establishment of the mutation spectrum by DNA sequencing. We describe various approaches for data analysis and interpretation of the results. Furthermore, we highlight representative studies in which the Big Blue® mouse cell culture model and the λ Select cII assay have been used for mutagenicity testing of diverse carcinogens. We delineate the advantages of this approach and discuss its limitations, while underscoring auxiliary methods, where applicable.
Highlights
The contemporary field of mutation research has witnessed a shift of interest from in vivo to in vitro mutagenicity testing to meet the requirements of “reducing” the number of animals used, “refining” procedures to alleviate animal pain, suffering or distress and “replacing” animal use with viable alternatives
Mutation assays in cultured cells of transgenic rodents constitute, in many ways, feasible “replacements” for in vivo mutagenicity experiments in the corresponding animals [2]
If used properly and in well-designed experiments, the in vitro models can provide an initial indication of the mutagenic potential of a given chemical/agent(s), and the results can be used as a guide to devise ‘refined’ in vivo experiments with a “reduced” number of animals [2,4]
Summary
The contemporary field of mutation research has witnessed a shift of interest from in vivo to in vitro mutagenicity testing to meet the requirements of “reducing” the number of animals used, “refining” procedures to alleviate animal pain, suffering or distress and “replacing” animal use with viable alternatives. Mutation assays in cultured cells of transgenic rodents constitute, in many ways, feasible “replacements” for in vivo mutagenicity experiments in the corresponding animals [2] These in vitro models offer significant advantages over their counterparts in vivo animal models as they are much less costly and laborious, require far less time to be completed and, most importantly, do not involve direct use of the animals [3,4,5]. If used properly and in well-designed experiments, the in vitro models can provide an initial indication of the mutagenic potential of a given chemical/agent(s), and the results can be used as a guide to devise ‘refined’ in vivo experiments with a “reduced” number of animals [2,4]
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