The majority of patients with multiple myeloma (MM) have persistence of minimal residual disease (MRD), as determined by polymerase chain reaction (PCR) detection of clonal immunoglobulin H (IgH) gene rearrangements. As a result, PCR analysis has not provided clinically useful prognostic information in myeloma patients. Instead, quantitative PCR approaches are required to predict patient outcomes and assess response to novel treatment strategies. We adapted real-time PCR technology to quantify myeloma cells using the IgH rearrangement and then assessed the utility of this approach in 29 patients with myeloma who had undergone autologous stem cell transplantation. Because of the high cost of producing a specific reporting probe for each patient, H-chain V-region family-specific consensus probes were used in association with allele-specific oligonucleotides for PCR amplification. Because of the high frequency with which somatic hypermutation at the immunoglobulin locus occurs in MM, a number of mismatches occurred between the patient sequences and the consensus probe. However, construction of a limited number of probes allowed real-time PCR with a sensitivity of 10(-4) to 10(-5). To validate this method, we extensively evaluated assay accuracy and reproducibility. Results indicate that real-time PCR using consensus probes provides a feasible, accurate, and reproducible method for evaluating MRD in M M and possibly in other differentiated B-cell malignancies, and one that is less expensive than the use of patient-specific probes. This technique is being used to assess tumor depletion after immunologic purging and changes in tumor burden in patients undergoing stem cell transplantation and novel treatment approaches.