Detection of monoclonal B-cell populations in decalcified, plastic-embedded bone marrow biopsies with the polymerase...

Polymerase chain reaction (PCR) has been employed successfully for the detection of clonal immunoglobulin gene rearrangements in paraffin-embedded clinical samples. The authors examined whether this technique can also be applied to fixed, decalcified, and plastic-embedded bone marrow biopsies. DNA extracted from 66 glycolmethacrylate-embedded trephine biopsy samples was amplified for the detection of rearranged VDJ regions of the immunoglobulin heavy chain genes using both a single-step and a semi-nested PCR technique. After exclusion of samples with inadequate DNA, clonality was confirmed in 16 (67%) of 24 cases with B cell malignancy, whereas all 11 non-B cell neoplasms, and 6 of 9 cases with normal bone marrow showed evidence of a polyclonal B cell population. Patterns indicating oligo- or monoclonality were observed in three plastic-embedded samples of normal bone marrow, although control PCR of frozen bone marrow samples obtained in parallel showed no evidence of clonality. Repeated PCR of these cases revealed inconsistent bands, probably due to the amplification of rare templates from polyclonal B cells. Decalcified, plastic-embedded bone marrow biopsies are suitable for PCR-based determination of B-cell clonality. To exclude the possibility of false-positive results, monitoring of template DNA quality and independent control amplifications are mandatory.