RNA quality and gene expression analysis of ovarian tumor tissue undergoing repeated thaw-freezing

Gene expression profiles evaluated by microarray-based quantization of RNA are used in studies of differential diagnosis and prognosis in cancer. RNA of good quality is mandatory for this evaluation. The RNA most often comes from tumor banks with limited amount of tissue, and the tissue often undergoes repeated thawing and freezing. We evaluated the influence of repeated division of tumor samples at room temperature, on RNA quality and quantity, in addition to the gene expression profile. Sixteen ovarian tumor samples were divided in three aliquots each, undergoing respectively one, two, and three thaw-freeze cycles. RNA from each aliquot was extracted on the day of division, and quantity and quality were evaluated. RNA from all three aliquots of four tumor samples underwent microarray analysis on Affymetrix Human Genome U133A 2.0 arrays. Microarray data were evaluated using both unsupervised, and supervised multivariate statistical methods, reliability analysis, as well as verification using published gene lists in ovarian cancer studies. RNA quality and quantity did not change during the division procedure and microarray data showed insignificant difference in gene expression. Tumor samples from tumor banks can be frozen and thawed at least three times without compromising the RNA integrity and genetic expression profile.