Heteroduplex PCR analysis of rearranged immunoglobulin genes for clonality assessment in multiple myeloma.

R. García-Sanz, R. López-Pérez, A.W. Langerak, D. González, M.C. Chillón, A. Balanzategui, M.V. Mateos, I. Alaejos, M. González, J.J. Van Dongen, J.F. San Miguel

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26 Citations (Scopus)

Abstract

BACKGROUND AND OBJECTIVE: Molecular analysis by PCR of monoclonally rearranged immunoglobulin (Ig) genes can be used for diagnosis in B-cell lymphoproliferative disorders (LPD), as well as for monitoring minimal residual disease (MRD) after treatment. This technique has the risk of false-positive results due to the "background" amplification of similar rearrangements derived from polyclonal B-cells. This problem can be resolved in advance by additional analyses that discern between polyclonal and monoclonal PCR products, such as the heteroduplex analysis. A second problem is that PCR frequently fails to amplify the junction regions, mainly due to somatic mutations frequently present in mature (post-follicular) B-cell lymphoproliferations. The use of additional targets (e.g. Ig light chain genes) can avoid this problem. DESIGN AND METHODS: We studied the specificity of heteroduplex PCR analysis of several Ig junction regions to detect monoclonal products in samples from 84 MM patients and 24 patients with B cell polyclonal disorders. RESULTS: Using two distinct VH consensus primers (FR3 and FR2) in combination with one JH primer, 79% of the MM displayed monoclonal products. The percentage of positive cases was increased by amplification of the Vlamda-Jlamda junction regions or kappa(de) rearrangements, using two or five pairs of consensus primers, respectively. After including these targets in the heteroduplex PCR analysis, 93% of MM cases displayed monoclonal products. None of the polyclonal samples analyzed resulted in monoclonal products. Dilution experiments showed that monoclonal rearrangements could be detected with a sensitivity of at least 10(-2) in a background with >30% polyclonal B-cells, the sensitivity increasing up to 10(-3) when the polyclonal background was
Original languageEnglish
Pages (from-to)328-335
Number of pages8
JournalHAEMATOLOGICA
Volume84
Issue number4
Publication statusPublished - Apr 1999

Keywords

  • B-Lymphocytes
  • Gene Rearrangement
  • Genes, Immunoglobulin
  • Humans
  • Multiple Myeloma
  • Polymerase Chain Reaction

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