Epithelial ovarian tumors frequently display deletions on the short arm of chromosome 3 suggesting the existence of tumor suppressor genes within the deleted regions. We have recently established a primary tissue culture system as a model to investigate the genetic events associated with ovarian cancer. The frequencies of loss of heterozygosity (LOH) at 16 loci representative of chromosome 3p in 33 tumor biopsies and 47 ovarian primary cultures derived from unselected ovarian cancers were examined. This repertoire also included benign and borderline tumors as well as malignant ovarian ascites. LOH was observed in 25 (31%) samples for at least one marker: 21 of 58 malignant, two of 12 borderline and two of 10 benign specimens. Chromosome 3p loss was not restricted to ovarian tumors of high grade and stage. LOH was observed in both cultured and non cultured tumors and ascites. A spontaneously immortalized cell line derived from a malignant ovarian ascites, OV-90, displayed LOH of the majority of markers suggesting loss of one homolog of chromosome 3p. The pattern of deletion displayed by these 25 samples enabled the determination of at least two distinct regions of overlapping deletions on chromosome 3p extending from D3S1270 to D3S1597 and from D3S1293 to D3S1283. In addition, a region proximal to D3S1300 was deleted in a subset of samples. Although loss of loci overlapping these three regions (Regions I, II and III) were observed in malignant and benign tumors, in borderline tumors loss was observed of markers representative of Region III only. While RARbeta is presently included in Region II, the minimal regions of deletion exclude VHL, TGFBR2, PTPase(gamma) and FHIT as candidate tumor suppressors in ovarian tumorigenesis.