Comparative analysis of interphase FISH and RT-PCR to detect bcr-abl translocation in chronic myelogenous leukemia and related disorders.
The t(9;22)(q34;q1 1) between the abl and bcr genes plays a pivotal role in the diagnosis and pathogenesis of chronic myelogenous leukemia (CML). Its detection is routinely accomplished by Southern blot analysis and karyotyping. Interphase fluorescence in situ hybridization (FISH) and reverse transcriptase-polymerase chain reaction (RT-PCR) are emerging molecular techniques that offer viable alternatives. We analyzed 40 samples of peripheral blood and bone marrow (CML, 16; acute myelogenous leukemia, 6; acute lymphoblastic leukemia [ALL], 1; chronic lymphoblastic leukemia, 2; myelodysplasias, 4; myeloproliferative disorders, unclassified, 3; nonleukemic hematologic malignancies, 3; hypercellular bone marrow, 1; normal control samples, 2; and K562 cell line samples, 2) for the presence of bcr-abl fusion gene and its messenger RNA (mRNA) transcript by FISH and RT-PCR, respectively. We compared the results with results of Southern blot analysis and karyotyping when available. Cost analysis was performed. Thirty-three samples were evaluable by FISH; 14 of 14 evaluable CML samples and one ALL sample were positive for bcr-abl by FISH (100%). The other 15 evaluable samples were negative; 16 of 16 (100%) and 13 of 16 (81%) of CML cases were positive for bcr-abl mRNA by RT-PCR (chemiluminescent blot method) and RT-PCR (colorimetric method), respectively. The ALL sample was positive by both RT-PCR methods. All other samples were negative by RT-PCR (chemiluminescent blot method), and all but 1 case of myeloproliferative disorder tested negative by RT-PCR (colorimetric method). We conclude the utility of FISH and RT-PCR is associated with certain limitations, such as insufficient RNA for RT-PCR and the occasional absence of internal positive FISH control signals. However, each procedure offers (with a high concordance rate) a specific and cost-effective alternative to Southern blot analysis and karyotyping and improved turnaround time for the detection of bcr-abl fusion gene or its mRNA transcript.
Tbakhi
A
A
Department of Clinical Pathology, Cleveland Clinic Foundation, OH 44195, USA.
Pettay
J
J
Sreenan
J J
JJ
Abdel-Razeq
H
H
Kalaycio
M
M
Hoeltge
G
G
Miller
M L
ML
Tubbs
R R
RR
D015139:Blotting, Southern / Q000191:economics; D001853:Bone Marrow / Q000737:chemistry; D002892:Chromosomes, Human, Pair 22; D002899:Chromosomes, Human, Pair 9; D003124:Colorimetry; D005456:Fluorescent Dyes / Q000032:analysis; D016044:Fusion Proteins, bcr-abl / Q000235:genetics*; D016313:Genes, abl / Q000235:genetics*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence / Q000191:economics; D007621:Karyotyping; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000235:genetics*; D007962:Leukocytes / Q000737:chemistry; D015513:Oncogene Proteins / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000191:economics; D011505:Protein-Tyrosine Kinases; D011518:Proto-Oncogene Proteins; D051562:Proto-Oncogene Proteins c-bcr; D012333:RNA, Messenger / Q000032:analysis; D012334:RNA, Neoplasm / Q000032:analysis; D014158:Transcription, Genetic; D014178:Translocation, Genetic
D005456:Fluorescent Dyes; D015513:Oncogene Proteins; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D011505:Protein-Tyrosine Kinases; D016044:Fusion Proteins, bcr-abl; C496613:BCR protein, human; D051562:Proto-Oncogene Proteins c-bcr