title: Real time polymerase chain reaction in diagnosis of chronic myeloid leukemia.
abstract: OBJECTIVE
  To compare the sensitivity and specificity of Real Time Polymerase Chain Reaction (RT-PCR) with conventional cytogenetics in diagnosis of chronic myeloid leukemia.
  
  
  METHODS
  A cross-sectional, analytical study.
  
  
  METHODS
  The Armed Forces Institute of Pathology (AFIP), Rawalpindi, from December 2010 to January 2012.
  
  
  METHODS
  A total number of 40 patients were studied, in which all were diagnosed as CML on peripheral blood and bone marrow aspiration. The subjects were tested for the presence of Philadelphia (Ph) chromosome by cytogenetics and BCR-ABL fusion gene by RT-PCR. 2-3 ml of venous blood was collected, half in sodium heparin (anti-coagulant) for cytogenetics and half in EDTA for PCR. For cytogenetics, cells were cultured for 72 hours in RPMI 1640 medium and examined by arresting in metaphase using Colchicine to identify Philadelphia chromosome. For PCR, RNA extraction was done by Tri Reagent LS (MRC, USA) and cDNA was synthesized using reverse transcriptase and gene specific primer. RT- PCR was done on ABI-7500. The positive samples were identified when fluorescence exceeded threshold limit. Results of cytogenetics and RT PCR were compared.
  
  
  RESULTS
  Out of the 40 patients, PCR showed 37 (92.5%) were positive and 3 (7.5%) were negative for BCR-ABL fusion gene, whereas in cytogenetics 28 (70%) were positive for Ph chromosome and 12 (30%) were negative for Ph chromosome. Sensitivity and specificity of cytogenetics was 75.6% and 100% respectively.
  
  
  CONCLUSIONS
  Real time PCR as compared to cytogenetics is less tedious, gives quick results, does not require multiple sampling due to culture failure and can be done on peripheral blood.
authors.affiliation: Department of Haematology, Armed Forces Institute of Pathology, Rawalpindi.
authors.forename: Sunila
authors.initials: S
authors.lastname: Tashfeen
authors.affiliation: Department of Haematology, Armed Forces Institute of Pathology, Rawalpindi.
authors.forename: Suhaib
authors.initials: S
authors.lastname: Ahmed
authors.affiliation: Department of Haematology, Armed Forces Institute of Pathology, Rawalpindi.
authors.forename: Farhat Abbas
authors.initials: FA
authors.lastname: Bhatti
authors.affiliation: Department of Haematology, Armed Forces Institute of Pathology, Rawalpindi.
authors.forename: Nadir
authors.initials: N
authors.lastname: Ali
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D014408:Biomarkers, Tumor / Q000097:blood*; D003430:Cross-Sectional Studies; D003582:Cytogenetics / Q000379:methods*; D005260:Female; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000175:diagnosis* / Q000235:genetics*; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D010677:Philadelphia Chromosome; D012333:RNA, Messenger; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D055815:Young Adult
chemical_list: D014408:Biomarkers, Tumor; D012333:RNA, Messenger