ISOLATION OF VIABLE HODGKIN AND REED-STERNBERG CELLS FROM HODGKIN-DISEASE TISSUES

Hodgkin disease (HD) is characterized by a small number of malignant H odgkin and Reed-Sternberg (H/RS) cells among a major population of non malignant cells. The analysis of H/RS cells has been hampered by their low frequency and fragility. Here, we describe the isolation of viabl e H/RS cells from HD affected tissues by high gradient magnetic cell s orting (MACS) according to expression of CD30. The cells were enriched to a purity of up to 50%, H/RS cells were distinguished from other CD 30(+) cells by the expression of CD15, their size and granularity, No CD30/CD15 double-positive cells could be enriched from a lymph node af fected by the lymphocyte predominant subtype of HD, activated lymph no des or peripheral blood of healthy donors. For two cases of HD individ ual MACS-purified H/RS cells and H/RS cells micromanipulated from tiss ue sections of the same lymphoma specimens were analyzed for Ig gene r earrangements. In both cases, identical V gene rearrangements were amp lified from both sources of H/RS cells, showing that H/RS cells were s uccessfully enriched. Moreover, the finding that in both cases no addi tional Ig gene rearrangements other than the ones identified in the H/ RS cells micromanipulated from tissue sections were amplified from the MACS-purified H/RS cells further supports the monoclonality of these cells throughout the affected lymph nodes. The isolation of viable H/R S cells ex vivo is prerequisite for a direct study of gene expression by those cells and of their interaction with cells in their vicinity.