LYMPHOCYTE PREDOMINANCE HODGKINS-DISEASE - LINEAGE AND CLONALITY DETERMINATION USING A SINGLE-CELL ASSAY

Lymphocyte predominance Hodgkin's disease (LPHD) is a clinically indol ent condition. Although there is evidence that the putative neoplastic cell in this disease, the ''L&H'' cell, is of B-cell lineage, there i s conflicting data concerning the clonality of these cells. Our study was aimed at clarifying the issue of lineage and clonality of the L&H cells of LPHD using a single-cell assay. Four cases of LPHD were studi ed. To circumvent the difficulties of obtaining fresh tissue and to be able to study representative cases, a new method was developed to obt ain single-cell suspensions of L&H cells from archival formalin-fixed paraffin-embedded tissue. Single L&H cells were identified by morpholo gy and immunostaining for epithelial membrane antigen, isolated using a micropipette, and subjected to polymerase chain reaction (PCR) ampli fication of the complematarity determining region 3 (CDR3) of the Ig h eavy chain (IgH) gene, which is B-cell clone-specific. The PCR product s were size-fractionated by polyacrylamide gel electrophoresis and rep resentative products were directly sequenced. Single T cells and small B cells were also isolated from the tissues and used as negative and positive controls, respectively. In all four cases of LPHD, the IgH CD R3 of single L&H cells could be amplified. Within each case, the IgH C DR3 of single L&H cells was found to be of different length or of diff erent sequence. Therefore, our results provide strong evidence for the B-cell origin of the L&H cells and the polyclonal nature of LPHD. (C) 1994 by The American Society of Hematology.