Molecular analysis of the immunoglobulin heavy chain gene in the diagnosis of primary cutaneous B cell lymphoma

The diagnosis of primary cutaneous B cell lymphoma can be difficult on the basis of histologic and immunophenotypic features alone. Previous polymerase chain reaction studies for detection of a clonal population in nodal B cell lymphomas have employed different primer pairs with detection sensitivities varying between 34% and 94% but there have been no comprehensive studies of primary cutaneous B cell lymphoma. We compared the sensitivity of different sets of consensus primers to amplify the CDR3 VDJ region of the immunoglobulin heavy chain gene in combination with an immunoglobulin heavy chain joining region consensus primer to detect a monoclonal population in 39 cases of primary cutaneous B cell lymphoma. Radiolabeled products were analyzed with denaturing 6% polyacrylamide gel electrophoresis. Sequence analysis was used to confirm amplification of clonal immunoglobulin heavy chain gene rearrangements and to establish whether somatic hypermutation can interfere with primer binding. Clonal immuno globulin heavy chain gene rearrangements were demonstrated in 79% of cases (74% with leader sequences, 64% with FR1, and 45% with FR3 primers). Somatic hypermutation at primer binding sites was confirmed in cases where a false negative result was obtained with the FR3 primer. Although monoplex polymerase chain reaction amplification using the leader sequence primers is the most sensitive method for detecting a clonal population, six primers are required in six different reactions. Our findings suggest initial analysis with the FR3 primer and subsequent analysis using leader sequences in negative cases. Our data indicate that the FR3 consensus primer alone is not sufficient for a comprehensive analysis of primary cutaneous B cell lymphoma.