title: Quantitation of bcl-2 oncogene in cultured lymphoma/leukemia cell lines and in primary leukemia B-cells by a highly sensitive RT-PCR method.
abstract: BACKGROUND
  The bcl-2 gene, isolated from the t(14;18) chromosomal translocation breakpoint, is able to prevent apoptotic death induced by various stimuli in different tissues. Therefore bcl-2 oncogene expression could be a key parameter for investigating the molecular mechanisms involved in the apoptosis of normal and neoplastic hematopoietic cells.
  
  
  METHODS
  In order to evaluate bcl-2 expression in both follicular B-lymphomas carrying or not carrying the 14;18 translocation and in lymphatic leukemias, we optimized an internal standard-based method of reverse transcriptase-polymerase chain reaction (RT-PCR) for the rapid quantitation of bcl-2 mRNA cellular levels. A simple purification of the reverse transcription products resulted in very high PCR efficiency, so that radioactive labelling of the amplification products was avoided.
  
  
  RESULTS
  bcl-2 mRNA levels proved to be higher in t(14;18) than in t(14;18) negative cell lines, and higher in primary leukemia pre-B cells than in early-B cells. Tested for sensitivity by identifying minimal residual t(14;18) B cells expressing the bcl-2/IgH gene, this RT-PCR method was able to detect bcl-2/IgH mRNA from just one t(14;18) positive cell out of ten million t(14;18) negative cells.
  
  
  CONCLUSIONS
  The RT-PCR method we optimized appears to be suitable for clinical use in both leukemia/lymphoma characterization and in lymphomatous disease follow-up.
authors.affiliation: Institute of General Pathology, University of Florence, Italy.
authors.forename: A
authors.initials: A
authors.lastname: Quattrone
authors.forename: L
authors.initials: L
authors.lastname: Papucci
authors.forename: V
authors.initials: V
authors.lastname: Santini
authors.forename: N
authors.initials: N
authors.lastname: Schiavone
authors.forename: D
authors.initials: D
authors.lastname: Noferini
authors.forename: A
authors.initials: A
authors.lastname: Calastretti
authors.forename: E
authors.initials: E
authors.lastname: Copreni
authors.forename: S
authors.initials: S
authors.lastname: Morelli
authors.forename: P L
authors.initials: PL
authors.lastname: Rossi Ferrini
authors.forename: A
authors.initials: A
authors.lastname: Nicolin
mesh_terms: D001483:Base Sequence; D015972:Gene Expression Regulation, Neoplastic / Q000502:physiology*; D006801:Humans; D015448:Leukemia, B-Cell / Q000235:genetics* / Q000473:pathology; D016393:Lymphoma, B-Cell / Q000235:genetics* / Q000473:pathology; D008969:Molecular Sequence Data; D009857:Oncogenes; D016133:Polymerase Chain Reaction / Q000379:methods; D012194:RNA-Directed DNA Polymerase; D014407:Tumor Cells, Cultured
chemical_list: D012194:RNA-Directed DNA Polymerase