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Published on: March 5, 2018
Quantitation of minimal residual disease in multiple myeloma using an allele-specific real-time PCR assay
T Rasmussen1, T S Poulsen, L Honoré
1Department of Hematology L, Herlev Hospital, University of Copenhagen, Herlev, Denmark. thra@herlevhosp.kbhamt.dk
Objective:
To develop a real-time PCR method, based on the 5'nuclease TaqMan technology, for quantitation of clonal cells in multiple myeloma (MM).
Materials And Methods:
The real-time quantitative PCR method incorporates both an allele-specific oligonucleotides (ASO) primer and an ASO dual-labeled fluorogenic probe (ASO TaqMan probe). The ASO primer and probe corresponded to the complementary determining region 3 (CDR3) of the rearranged immunoglobulin heavy chain gene (IgH). With the use of a sequence detector, PCR product accumulation was measured through the ASO TaqMan probe. The real-time PCR method was compared with flow cytometric quantitation of myeloma plasma cells.
Results:
The application of the real-time quantitative ASO IgH PCR method is illustrated by a sequential analysis of minimal residual disease (MRD) in bone marrow (BM) samples from myeloma patients undergoing peripheral blood stem cell (PBSC) transplantation. The real-time PCR method was able to quantitate residual malignant cells in BM samples from patients who were considered to be in complete remission. Further, it was illustrated that a potential problem in determining tumor cell content in myeloma BM samples is the heterogeneous infiltration of the marrow.
Conclusion:
The application of the real-time PCR method provides a sensitive, highly specific, and reproducible quantitation of myeloma cells.
