Molecular methods for the detection of mutations

We report the results of a collaborative study aimed at developing reliable , direct assays for mutation in human cells. The project used common lympho blastoid cell lines, both with and without mutagen treatment, as a shared r esource to validate the development of new molecular methods for the detect ion of low-level mutations in the presence of a large excess of normal alle les. As the "gold standard," hprt mutation frequencies were also measured o n the same samples. The methods under development included i) the restricti on site mutation (RSM) assay, in which mutations lead to the destruction of a restriction site; ii) minisatellite length-change mutation, in which mut ations lead to alleles containing new numbers of tandem repeat units; iii) loss of heterozygosity for HLA epitopes, in which antibodies can be used to direct selection for mutant cells; iv) multiple fluorescence-based long Li nker arm nucleotides assay (mf-LLA) technology, for the detection of substi tutional mutations; vi detection of alterations in the TP53 locus using a ( CA) array as the target for the screening: and vi) PCR analysis of lymphocy tes for the presence of the BCL2 t(14:18) translocation. The relative merit s of these molecular methods are discussed, and a comparison made with more "traditionaI" methods. Teratogenesis Carcinog. Mutagen. 20:357-386. 2000. (C) 2000 Wiley-Liss, Inc.