The aim of this study was to find out whether UICC crocidolite asbestos fibers induce in vivo DNA double-strand breaks in the nuclei of cells in lung and liver of rats. The effect of crocidolite on chromosomal DNA of rats exposed to crocidolite fibers was analyzed by pulsed-field gel electrophoresis (PFGE) using the chromosomal DNA-protective agarose-plugs method and laser densitometric evaluation of stained DNA bands. The profiles of laser densitometric scanning showing PFGE separation of DNA for exposed rats are right-shifted in liver and lung compared with the controls of the same age. DNA from the liver of nonexposed rats showed one peak in the range of 250–850 kb. Compared to this finding, at 4, 13, and 16 mo after crocidolite instillation DNA of rats showed shorter DNA fragments in the range of 250–700 kb. This indicates a more intact DNA in controls. At 4, 13, and 16 mo after crocidolite instillation, we also found DNA fragments in exposed lungs in the region between 250 and 2200 kb produced by multiple double-strand breaks at random sites of the same rats. At 4 and 13 mo after instillation, DNA double-strand breaks were mainly found in asbestos lungs and only partly in para-asbestos lungs. No DNA double-strand breaks could be observed in the opposite lung (contralateral lung). At 76 mo after crocidolite instillation, DNA double-strand breaks could be detected in all parts of the lungs. It should be pointed out that occupational crocidolite exposure could be associated with DNA damage in lung and liver nuclei. The increased incidence of double-strand DNA fragments can be an important factor in the chronic effect of crocidolite asbestos-associated carcinogenesis.