title: Development of an anti-porcine CD34 monoclonal antibody that identifies hematopoietic stem cells.
abstract: OBJECTIVE
  The isolation of porcine hematopoietic stem cells (HSC) would be an important step toward development of porcine-to-human chimerism for induction of tolerance in clinical xenotransplantation. CD34 is a common marker of HSC and has not been developed as a marker in pigs. In this study we have generated and characterized a monoclonal antibody (mAb) that identifies porcine CD34 on a subset of porcine bone marrow (BM) stem/progenitor cells.
  
  
  METHODS
  The porcine CD34 gene was cloned and a recombinant protein produced. An anti-porcine CD34 mAb was produced that could detect both the recombinant protein and a subset of porcine BM cells. The CD34(+) cells were phenotyped by lineage and HSC associated markers. Furthermore, the CD34(+) cells were analyzed by colony-forming unit (CFU) assay.
  
  
  RESULTS
  Two splice variants of the porcine CD34 gene were cloned and a recombinant protein produced for mAb production. The mAb developed can detect both the recombinant protein and the native CD34 protein on a range of pig tissues, including BM. This subset of BM cells was negative for hematopoietic lineage makers, including CD3, CD14, and CD21 and positive for other known porcine HSC markers, including CD90, CD172a, histocompatibility complex (MHC) class I, and MHC class II. Moreover, the CD34(+) BM cells were enriched for multilineage progenitor cells as determined by CFU assay.
  
  
  CONCLUSIONS
  Similar to human and mouse CD34, pig CD34 detects a subset of BM progenitor cells. This mAb will now provide a means for isolating porcine CD34(+) cells to be further analyzed for HSC activity and to assess their potential to develop pig-to-human chimeras to induce xenograft tolerance.
authors.affiliation: CSIRO Livestock Industries, Australian Animal Health Laboratories, Geelong, Victoria, Australia. daniel.layton@csiro.au
authors.forename: Daniel S
authors.initials: DS
authors.lastname: Layton
authors.forename: A David G
authors.initials: AD
authors.lastname: Strom
authors.forename: Terri E
authors.initials: TE
authors.lastname: O'Neil
authors.forename: Mary M
authors.initials: MM
authors.lastname: Broadway
authors.forename: Garth L
authors.initials: GL
authors.lastname: Stephenson
authors.forename: Kirsten R
authors.initials: KR
authors.lastname: Morris
authors.forename: Morley
authors.initials: M
authors.lastname: Muralitharan
authors.forename: Mauro S
authors.initials: MS
authors.lastname: Sandrin
authors.forename: Francesco L
authors.initials: FL
authors.lastname: Ierino
authors.forename: Andrew G D
authors.initials: AG
authors.lastname: Bean
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal; D018952:Antigens, CD34 / Q000235:genetics / Q000276:immunology*; D001854:Bone Marrow Cells; D018929:Cell Culture Techniques; D002469:Cell Separation / Q000379:methods*; D003001:Cloning, Molecular; D003114:Colony-Forming Units Assay; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000276:immunology*; D016130:Immunophenotyping / Q000379:methods; D013552:Swine
chemical_list: D000911:Antibodies, Monoclonal; D018952:Antigens, CD34