title: Generation of anti-human DEC205/CD205 monoclonal antibodies that recognize epitopes conserved in different mammals.
abstract: DEC205/CD205 is a C-type multilectin receptor, expressed highly in dendritic cells (DCs). Previous efforts to generate anti-human DEC205 (anti-hDEC205) monoclonal antibodies (mAbs) from mice immunized with subdomain proteins of hDEC205 resulted in a few mAbs. Recently, we expressed and utilized a full-length extracellular domain protein of hDEC205 to successfully generate 5 strong anti-hDEC205 mAbs from mice. In this study, DEC205 knockout (KO) mice were immunized with this full-length extracellular domain protein of hDEC205. One of the 3 immunized DEC205 KO mice was chosen for the highest anti-hDEC205 titer by flow cytometric analysis of serum samples on CHO cells stably expressing hDEC205 (CHO/hDEC205 cells) and used for hybridoma fusion. From a single fusion, more than 400 anti-hDEC205 hybridomas were identified by flow cytometric screen with CHO/hDEC205 cells, and a total of 115 hybridomas secreting strong anti-hDEC205 mAb were saved and named HD1 through HD115. To characterize in detail, 10 HD mAbs were chosen for superior anti-hDEC205 reactivity and further subjected to cloning and purification. Interestingly, out of those 10 chosen anti-hDEC205 HD mAbs, 5 mAbs were also strongly reactive to mouse DEC205 while 8 mAbs were found to stain DEC205(+) DCs on monkey spleen sections. In addition, we also identified that HD83, one of the 10 chosen HD mAbs, stains DEC205(+) DCs in rat spleen and lymph node. Therefore, by immunizing DEC205 KO mice with a full-length extracellular domain protein of hDEC205, we generated a large number of strong anti-hDEC205 mAbs many of which are cross-species reactive and able to visualize DEC205(+) DCs in lymphoid tissues of other mammals.
authors.affiliation: Laboratory of Cellular Physiology and Immunology and Chris Browne Center for Immunology and Immune Diseases, The Rockefeller University, New York, NY 10065, USA. parkc@rockefeller.edu
authors.forename: Chae Gyu
authors.initials: CG
authors.lastname: Park
authors.forename: Anthony
authors.initials: A
authors.lastname: Rodriguez
authors.forename: Hisashi
authors.initials: H
authors.lastname: Ueta
authors.forename: Haekyung
authors.initials: H
authors.lastname: Lee
authors.forename: Maggi
authors.initials: M
authors.lastname: Pack
authors.forename: Kenjiro
authors.initials: K
authors.lastname: Matsuno
authors.forename: Ralph M
authors.initials: RM
authors.lastname: Steinman
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000276:immunology; D015703:Antigens, CD / Q000276:immunology*; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D003713:Dendritic Cells / Q000276:immunology*; D000939:Epitopes / Q000276:immunology; D006801:Humans; D007150:Immunohistochemistry; D037181:Lectins, C-Type / Q000276:immunology*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D015778:Minor Histocompatibility Antigens; D011956:Receptors, Cell Surface / Q000276:immunology*; D011993:Recombinant Fusion Proteins / Q000276:immunology; D013047:Specific Pathogen-Free Organisms; D014162:Transfection
chemical_list: D000911:Antibodies, Monoclonal; D015703:Antigens, CD; C093569:DEC-205 receptor; D000939:Epitopes; D037181:Lectins, C-Type; D015778:Minor Histocompatibility Antigens; D011956:Receptors, Cell Surface; D011993:Recombinant Fusion Proteins