Nonradioactive detection of monoclonal immunoglobulin heavy chain gene rearrangement with PCR-SSCP.

Abstract

The rearrangement of the immunoglobulin heavy chain gene can be used as a marker of B-cell lineage and clonality. By using polymerase chain reaction (PCR) with variable-region and joining-region specific primers it is possible to detect the rearrangement of a small amount of clonal B cells, as described by several groups. The specific PCR product can be detected after amplification with gel electrophoresis and ethidium bromide staining. The DNA fragments obtained from different clones are, however, approximately of the same size, making it difficult to distinguish between the clones by simple electrophoresis and ethidium bromide staining, as described in many reports. Single-strand conformational polymorphism (SSCP) was evaluated as a method to detect specific clonal amplicons in a mixture of PCR-amplified products. Unique patterns were obtained for different B-cell clones, detectable in mixtures of 0.25% clonal cells in normal cells. It is concluded that SSCP is a valuable method for the specificity control of PCR in B-lymphocyte clonality analyses. The advantages of the described method over previously published techniques are increased specificity, simplicity without radioactivity, and rapidity.