Real-time 5' → 3' exonuclease-based PCR assay for detection of the t(11;14)(q13;q32)

• Published in: American journal of clinical pathology Vol. 112; no. 4; pp. 524 - 530
• Main Authors: LUTHRA, R, SARRIS, A. H, HAI, S, PALADUGU, A. V, ROMAGUERA, J. E, CABANILLAS, F. F, MEDEIROS, L. J
• Format: Conference Proceeding Journal Article
• Language: English
• Published: Chicago, IL American Society of Clinical Pathologists 01.10.1999
• Subjects: Biological and medical sciences; Chromosomes, Human, Pair 11; Chromosomes, Human, Pair 14; Computer Systems; Exodeoxyribonuclease V; Exodeoxyribonucleases - metabolism; Hematology; Humans; Immunoglobulin Heavy Chains - genetics; Immunoglobulin Joining Region - genetics; Investigative techniques, diagnostic techniques (general aspects); Lymphoma, Non-Hodgkin - genetics; Medical sciences; Pathology. Cytology. Biochemistry. Spectrometry. Miscellaneous investigative techniques; Polymerase Chain Reaction - methods; Taq Polymerase - metabolism; Translocation, Genetic; Chromosomal aberration; Human; Enzyme; Esterases; Malignant hemopathy; B-Lymphocyte; Phosphoric diester hydrolases; Non Hodgkin lymphoma; Real time; Polymerase chain reaction; Lymphoproliferative syndrome; Mantle cell lymphoma; Chromosome 11; Abnormal chromosome; Hydrolases; Chromosome 14; Diagnosis; Technique; Molecular biology; Phosphodiesterase I; Chromosome translocation
• ISSN: 0002-9173; 1943-7722
• DOI: 10.1093/ajcp/112.4.524

We describe the usefulness of a real-time polymerase chain reaction (PCR) assay for detection of the t(11;14)(q13;q32), most commonly present in mantle cell lymphoma (MCL). This assay is based on the 5'-->3' exonuclease activity of Taq polymerase, which cleaves an internal probe labeled with a reporter dye at its 5' end and a quencher dye at its 3' end during PCR. The real-time t(11;14) PCR assay was established using DNA from a case of MCL with the t(11;14), amplifiable using conventional PCR and primers specific for the major translocation cluster (MTC) region of the bcl-1 locus and the immunoglobulin heavy chain joining region gene (JH). The specificity was determined by analyzing DNA from 82 cases: 50 MCL, 27 other types of non-Hodgkin lymphoma (NHL), and 5 reactive lymphoid proliferations. The real-time t(11;14) PCR results were correlated with data obtained by a conventional PCR assay. By using the real-time assay, bcl-1 MTC/JH DNA fusion sequences were detected in 25 of 50 MCLs but not in other NHLs or reactive lymphoid proliferations. Concordance between real-time and conventional PCR methods for MCL was 96% and for all samples was 98%. The results demonstrate that this real-time PCR method to detect bcl-1 MTC/JH DNA fusion sequences is specific and reliable. In addition, the results are available immediately following amplification, without standard post-PCR manipulations.