Molecular diagnostic testing for determination of myeloid lineage in acute leukemias.
Determination of myeloid vs. lymphoid cell lineage in acute leukemias is essential for diagnosis, prognosis, and treatment. However, some leukemic cells are too primitive to be identified based on conventional morphology, cytochemistry, and immunophenotype criteria. The gene for myeloperoxidase (MPO) is central to the lineage designation of myeloid cells and their function. Expression of MPO messenger ribonucleic acid (mRNA) may serve to determine the myeloid lineage of primitive leukemic cells that do not express the final product, MPO. Recently, a procedure has been developed for detection of MPO mRNA in leukemic cells, based on message amplification by reverse transcription-polymerase chain reaction (RT-PCR). Three variant polymerase chain reaction (PCR) methods, developed in view of future diagnostic applications of the RT-PCR procedure are reported: (A) nested PCR, using a second pair of internal primers for reamplification of the first PCR product; (B) multiplex PCR for concomitant amplification of up to seven sequences encompassing all regions of MPO mRNA; (C) adaptation of the standard PCR procedure and the two variant methods for use of cellular material from air-dried unstained or Wright stained smears. These variant procedures developed on the HL-60 cell line make the RT-PCR detection of MPO mRNA easily applicable as a new diagnostic test in acute leukemias and confer the versatility needed in the clinical setting. The advantages of our new procedure for MPO mRNA detection are: high sensitivity and specificity, speed, simplicity, and safety. Myeloid lineage of leukemic blasts can be established, confirmed, or ruled out in 6 to 10 hours, making results available at the time of bone marrow sign-out.(ABSTRACT TRUNCATED AT 250 WORDS)
Crisan
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Department of Clinical Pathology, William Beaumont Hospital, Royal Oak, MI 48073.
D000199:Actins / Q000235:genetics; D000208:Acute Disease; D001483:Base Sequence; D015139:Blotting, Southern; D018076:DNA, Complementary / Q000235:genetics; D006801:Humans; D007938:Leukemia / Q000473:pathology*; D015335:Molecular Probes / Q000235:genetics; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D009195:Peroxidase / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods; D012333:RNA, Messenger / Q000378:metabolism; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
D000199:Actins; D018076:DNA, Complementary; D015335:Molecular Probes; D012333:RNA, Messenger; D009195:Peroxidase