Ovarian cancer, as well as other cancers, is primarily caused by methylation at cytosines in CpG islands, but the current marker for ovarian cancer is low in sensitivity and failed in early-stage detection. Fourier transform infrared (FT-IR) spectroscopy is powerful in analysis of functional groups within molecules, and infrared microscopy illustrates the location of specific groups within single cells. In this study, we applied HPLC and FT-IR microspectrometry to study normal epithelial ovarian cell line immortalized ovarian surface epithelium (IOSE), two epithelial ovarian cell lines (A2780 and CP70) with distinct properties, and the effect of a cancer drug 5-aza-2'-deoxycytidine (5-aza) without labeling. Our results reveal that inhibition of methylation on cytosine with 5-aza initiates the protein expression. Furthermore, paraffin-adsorption kinetic study allows us to distinguish hypermethylated and hypomethyated cells, and this assay can be a potential diagnosis method for cancer screening.