Molecular cloning and characterization of markers and cytokines for equid myeloid cells Falko Steinbach a, * , Robert Stark b , Sherif Ibrahim a , Eman Abd-El Gawad a , Hanns Ludwig c , Jakob Walter d , Ulrich Commandeur e , Susanne Mauel c a Institute for Zoo and Wildlife Research, Alfred-Kowalke-Str. 17, 10315 Berlin, Germany b Institute of Veterinary-Pathology, FU Berlin, Robert von Ostertag Str. 15, 14163 Berlin, Germany c Institute of Virology, FU Berlin, Ko ¨nigin-Luise-Str. 49, 14195 Berlin, Germany d Experimental Toxicology, Schering AG, Mu ¨llerstrasse 178, 13342 Berlin, Germany e Institute of Molecular Biotechnology, RWTH Aachen, Worringer Weg 1, 52074 Aachen, Germany Abstract The myeloid cell system comprises of monocytes, macrophages (MF), dendritic cells (DC), Kupffer cells, osteoclasts or microglia and is also known as the mononuclear phagocytic system (MPS). Essential cytokines to differentiate or activate these cells include GM-CSF or IL-4. Important markers for characterization include CD1, CD14, CD68, CD163 and CD206. All these markers, however, were not cloned or further characterized in equids by use of monoclonal antibodies earlier. To overcome this problem with the present study, two approaches were used. First, we cloned equine cytokines and markers, and second we analyzed cross-reactivity of human homologues or anti-human monoclonal antibodies. For cloning of equine cytokines and markers, we used degenerate primers delineated from other species, or equine-specific primers based on previous information in Genbank. Flow cytometry was used to determine the expression of markers on myeloid cells. Cross-reactivity could be shown for anti-human CD14, CD163 and mannose receptor (CD206) mAbs. Surface markers such as CD1 and CD68 that distinguish MF and DC were cloned and sequenced. According to blast homology, equine CD1a and CD1b could be identified and distinguished. With the resulting information, dendritic cells and macrophages of horses may be characterized. # 2005 Elsevier B.V. All rights reserved. Keywords: CD; Cloning; Cytokines; Equine; Monocyte; Macrophage 1. Introduction Cells that derive from pro-monocytes/monocytes are grouped in the mononuclear phagocytic system (MPS), although many of them are not professional phagocytes. Various types of macrophages (MF), such as alveolar MF or soft tissue histiocytes, microglia, Kupffer cells, and dendritic cells (DC) www.elsevier.com/locate/vetimm Veterinary Immunology and Immunopathology 108 (2005) 227–236 * Corresponding author. Present address: Virology Department, Veterinary Laboratories Agency, Woodham Lane, New Haw, Addlestone, Surrey KT15 3NB, UK. Tel.: +44 1932 357 566; fax: +44 1932 357 239. E-mail address: f.steinbach@vla.defra.gsi.gov.nk (F. Steinbach). 0165-2427/$ – see front matter # 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.vetimm.2005.07.015