The placenta plays an essential role in fetal growth and the maintenance of pregnancy and its functions are strictly controlled in a stage-specific manner. To gain an insight into placental functions and their regulation, we analyzed the plasma membrane proteins of rat placenta by two-dimensional polyacrylamide gel electrophoresis (2D/E). Plasma membrane fractions of the placenta obtained on days 12, 14, 16, 18 and 20 of pregnancy were purified by Percoll gradient centrifugation, and subjected to 2D/E analysis. After the proteins on the 2D/E gels had been visualized by silver staining, the patterns on the gels at different stages of pregnancy were compared using image analysis software. Proteins within an isoelectric point (pI) range of 4.0 to 7.0 and a molecular weight (Mw) range of 20-100 kDa were analyzed in detail, and about 800 proteins on average were recognized on each gel. Of these, the expression of 150 proteins was found to change dramatically according to the stage of pregnancy. According to their expression patterns, these proteins were categorized into two groups, Group I and Group II. The proteins belonging to Group I showed a higher intensity of expression on day 12 and disappeared on day 20. They included 119 plasma membrane proteins and were divided into five subgroups. Group II, which consisted of three subgroups, included 31 proteins showing a low or negligible expression on day 12 and higher expression on day 20. Most of the other membrane proteins (about 600) were expressed constantly during pregnancy. On the basis of our data, we constructed a database for plasma membrane proteins of the rat placenta.