We have investigated previously the utility of oligonucleotide expression microarray technology in an analysis of four spontaneously transformed epithelial ovarian cancer (EOC) cell lines, TOV-21G, TOV-81D, OV-90, and TOV-112D. Here, we examine the expression of 290 expressed sequence tags (ESTs) that map to human chromosome 3 in a primary culture derived from normal ovarian surface epithelium (NOSE), NOV-31, and the four spontaneously transformed EOC cell lines. One of these cell lines, OV-90, harbors a deletion of an entire chromosome 3p arm. Whereas the most aggressive cell lines (OV-90, TOV-112D, and TOV-21G) exhibited the highest levels of expression, assessed by the mean of expression values of all ESTs, OV-90 showed the lowest mean of expression of ESTs that map to the 3p arm in comparison with TOV-112D and TOV-21G. This difference in expression profile of 3p ESTs in OV-90 is also reflected in the ratio of expression of ESTs on 3p versus the 3q arm and in that the expression values of ESTs that map to 3p were more often lower than higher in OV-90 in two-way comparisons with NOV-31, TOV-21G, and TOV-112D. The loss of a 3p arm does not affect the pattern of differential expression in analyses based on the range of numeric expression values of each EST or fold differences in expression for each EST in comparison with NOV-31. However, 25 differentially expressed ESTs were identified on the basis of threefold differences in expression values between NOV-31 and any EOC cell line; and six of these ESTs were differentially expressed uniquely in OV-90. The investigation of these ESTs could facilitate the identification of novel chromosome 3 genes implicated in ovarian tumorigenesis.