title: Detection of tumor cells in purged bone marrow and peripheral-blood mononuclear cells by polymerase chain reaction amplification of bcl-2 translocations.
abstract: OBJECTIVE
  To compare bone marrow (BM) before and after purging with monoclonal antibodies (MAbs) and complement with peripheral-blood mononuclear cells (PBMNCs) for tumor-cell contamination by amplification of t(14;18) sequences using the polymerase chain reaction (PCR).
  
  
  METHODS
  Sixty patients with non-Hodgkin's lymphoma (NHL) undergoing autologous BM transplantation were evaluated. Six BM biopsies were performed at the time of harvesting and evaluated morphologically for tumor involvement. The harvested BM was treated with a panel of anti-B-cell MAbs directed against CD9, CD10, CD19, and CD20, followed by rabbit complement. Clonogenic assays were performed before and after purging. DNA was extracted and t(14;18) sequences amplified by PCR. PBMNCs collected by apheresis for back-up purposes were similarly evaluated.
  
  
  RESULTS
  Fifteen patients (25%) were PCR-positive before BM purging. Following MAb- and complement-mediated purging, there was a reduction in the PCR-amplified signal in 10 patients (67%). There was no reduction in colony-forming unit granulocyte-macrophage (CFU-GM) colony growth following purging. Eight of these 15 patients (53%) had morphologic evidence of BM involvement at the time of harvesting. In these eight patients, only three had a reduction in the PCR-amplified products, as compared with all seven who were morphologically negative at the time of BM harvesting (P = .026). Fourteen of these 15 patients had PBMNCs collected near the time of BM harvesting and 12 (86%) were PCR-positive.
  
  
  CONCLUSIONS
  BM purging with MAbs and complement results in reduction in the number of t(14;18)-positive tumor cells, especially in those patients who have no morphologic evidence of BM disease at the time of harvesting. Purged BM was less contaminated with t(14;18)-positive cells than unpurged PBMNCs, which were frequently contaminated with tumor cells.
authors.affiliation: Division of Hematology, Stanford University Medical Center, CA.
authors.forename: R S
authors.initials: RS
authors.lastname: Negrin
authors.forename: J
authors.initials: J
authors.lastname: Pesando
mesh_terms: D000293:Adolescent; D000328:Adult; D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D001483:Base Sequence; D001853:Bone Marrow / Q000276:immunology / Q000473:pathology* / Q000648:ultrastructure; D001856:Bone Marrow Examination; D016465:Bone Marrow Purging; D016026:Bone Marrow Transplantation; D002648:Child; D002883:Chromosomes, Human, Pair 14; D002887:Chromosomes, Human, Pair 18; D003114:Colony-Forming Units Assay; D003165:Complement System Proteins; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000276:immunology / Q000473:pathology* / Q000648:ultrastructure; D008228:Lymphoma, Non-Hodgkin / Q000235:genetics / Q000276:immunology / Q000473:pathology* / Q000628:therapy*; D008875:Middle Aged; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011518:Proto-Oncogene Proteins / Q000032:analysis*; D019253:Proto-Oncogene Proteins c-bcl-2; D014178:Translocation, Genetic
chemical_list: D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D003165:Complement System Proteins