The development of human breast cancer is a complex multi-step process that depends on various exogenous and endogenous factors that modulate the transformation of normal human breast epithelial cells into neoplastic ones. Using a spontaneously-immortalized human breast epithelial (MCF-10F) cell line, we have shown previously that radiation, in combination with estrogen, induces a stepwise neoplastic transformation of this cell line. In the present study, we investigate the incidence of microsatellite instability and loss of heterozygosity using a battery of markers on chromosomes 6 and 17, we correlate the genetic alteration with the malignant transformation of the MCF-10F cell line ranging from altered morphology to increase in proliferative rate, anchorage independent growth and tumorigenicity in nude mice. Microsatellite markers were selected from the hot spot regions (6q21-q27, 17p12-p13.3 and 17q12-q21) of both chromosomes. We found that the frequency of allelic imbalance occurs at the different stages of tumor progression with a range of 21 to 50% depending on the marker studied. The relatively high rate of allele imbalance at all these loci suggests the presence and inactivation of one or more tumor suppressor genes in these regions. Thus, the present data will be useful for systematic studies to identify the cellular and molecular changes associated with radiation-induced breast carcinogenesis.