Lipids are an energy source and key components of the cell membrane; however, they are also bioactive mediators of physiological and pathophysiological phenomena. Quantification of bioactive lipids is not easy because they have diverse chemical properties and are present in trace amounts. Here, we improved a multiplex method of quantifying bioactive lipids, thereby enabling measurement of 90 compounds simultaneously. We then used this system to quantify bioactive lipids produced by two subsets of dendritic cells (DCs): all-trans retinoic acid -treated DCs (RA-DCs) (a type of tolerogenic DC (tDC)) and conventional DCs (cDCs). We found that cDCs produced inflammatory lipid mediators such as leukotrienes, whereas RA-DCs produced anti-inflammatory lipid mediators such as prostaglandin 12. Consistent with this, cDCs expressed larger amounts of mRNA encoding 5-lipoxygenase and LTA4 hydrolase (both responsible for leukotriene biosynthesis) and RA-DCs produced larger amounts of mRNA encoding prostaglandin 12 synthase. Taken together, the results suggest that the method is useful for clarifying the roles of bioactive lipids during immune responses. (C) 2018 Elsevier Inc. All rights reserved.