Generation and Characterization of Anti-CD34 Monoclonal Antibodies that React with Hematopoietic Stem Cells

Cd34 is a type i membrane protein with a molecular mass of approximately 110 kda.this antigen is associated with human hematopoietic progenitor cells and is a differentiationstage-specific leukocyte antigen. in this study we have generated and characterizedmonoclonal antibodies (mabs) directed against a cd34 marker. mice wereimmunized with two keyhole lympet hemocyanin (klh)-conjugated cd34 peptides.fused cells were grown in hypoxanthine, aminopterine and thymidine (hat) selectivemedium and cloned by the limiting dilution (l.d) method. several monoclones wereisolated by three rounds of limited dilutions. from these, we chose stable clones thatpresented sustained antibody production for subsequent characterization. antibodieswere tested for their reactivity and specificity to recognize the cd34 peptides and furtherscreened by enzyme-linked immunosorbent assay (elisa) and western blottinganalyses. one of the mabs (3d5) was strongly reactive against the cd34 peptide andwith native cd34 from human umbilical cord blood cells (ucb) in elisa and westernblotting analyses. the results have shown that this antibody is highly specific andfunctional in biomedical applications such as elisa and western blot assays. thismonoclonal antibodies (mab) can be a useful tool for isolation and purification of humanhematopoietic stem cells (hscs).