title: A novel method to isolate and map endothelial membrane proteins from pulmonary vasculature.
abstract: Vascular endothelium has attracted extensive attention due to its important role in many physiological and pathological processes. Many methods have been developed to study the components and their functions in vascular endothelium. Here we report a novel approach to investigate vascular endothelium using normal rat lungs as the model. We perfused lung vascular beds with sulfosuccinimidyl-6-(biotinamido) hexanoate, a biotin analog, to label endothelial membrane proteins. The biotinylated proteins were isolated from lung homogenate with immobilized monomeric avidin and confirmed to be highly pure endothelial membrane proteins with little contamination of intracellular proteins. These biotinylated proteins were used as immunogens for development of monoclonal antibodies. Indeed, newly generated monoclonal antibodies have revealed different expression patterns of proteins across tissues. Some proteins were found highly specifically expressed to capillary vessels of pulmonary vasculature. This method has also been proven useful for investigating vasculature of other organs, as this study explored.
authors.affiliation: Institute of Molecular Medicine and State Key Laboratory of Pharmaceutical Biotechnology, Nanjing Univ., 22 Hankou Rd., Nanjing 210093, China.
authors.forename: Ying-Jiang
authors.initials: YJ
authors.lastname: Zhou
authors.forename: Shi-Quan
authors.initials: SQ
authors.lastname: Wang
authors.forename: Jing
authors.initials: J
authors.lastname: Zhang
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.forename: Feng
authors.initials: F
authors.lastname: Bi
authors.forename: Zhi-Gang
authors.initials: ZG
authors.lastname: Guo
authors.forename: Bi-Sen
authors.initials: BS
authors.lastname: Ding
authors.forename: Pat
authors.initials: P
authors.lastname: Kumar
authors.forename: Jian-Ning
authors.initials: JN
authors.lastname: Liu
authors.forename: Xiang-Yang
authors.initials: XY
authors.lastname: Tan
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D019921:Biotinylation / Q000379:methods*; D002462:Cell Membrane / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D002846:Chromatography, Affinity; D004730:Endothelium, Vascular / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D005260:Female; D015151:Immunoblotting; D007150:Immunohistochemistry; D008168:Lung / Q000098:blood supply*; D008565:Membrane Proteins / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D000911:Antibodies, Monoclonal; D008565:Membrane Proteins