Proteomic Analysis of Macrophages: a New Way to Identify Novel Cell-surface Antigens

Macrophages are involved in many important biological processes and membrane proteins are the key effector molecules for their function. However, membrane proteins are difficult to analyze by 2-DE based methods because of their intrinsic tendency to self-aggregate during the first dimension separation (IEF). To circumvent this, we combined one-dimensional SDS-PAGE with capillary liquid chromatography-tandem mass spectrometry (LC-MS/MS). Using this technique, we identified 458 GO annotated membrane proteins with extremely high confidence, including most known markers of peritoneal macrophages (e.g., CD11b, F4/80, CD14, CD18, CD86, CD44, CD16 and Toll-like receptor). Thirteen other CD antigens (CD243, CD98, CD107a, CD107b, CD36, CD97, CD205, CD206, CD180, CD191, CD300, CD45and CD29), and 18 Ras-related small GTPases were also identified. In addition to those known macrophage membrane proteins, a significant number of novel proteins have also been identified. This research not only provides a technique to study membrane proteins, but also a valuable dataset of macrophage antigens, thus providing better understanding of the functional mechanisms of macrophages in many biological processes.