IDENTIFICATION AND STRUCTURAL-ANALYSIS OF REARRANGED IMMUNOGLOBULIN HEAVY-CHAIN GENES IN LYMPHOMAS AND LEUKEMIAS

The third complementarity determining region (CDR3) of the hypervariab le domain of immunoglobulin heavy chain (IgH) genes represents a highl y variable and clone-specific region within the rearranged IgH gene lo cus. We used a polymerase chain reaction (PCR) assay to characterize d one-specific IgH-CDR3 sequences in 10 non-Hodgkin's lymphomas (NHL), f ive chronic lymphocytic leukemias (CLL) and five acute lymphoblastic l eukemias (ALL) of B cell lineage. The IgH-CDR3 sequences were amplifie d using DNA extracted from clinical specimens (bone marrow, peripheral blood and fresh-frozen or paraffin-embedded lymph nodes) by a semi-ne sted PCR with consensus primers directed to conserved regions within t he variable (V-H) and the joining (J(H)) gene segments. In 17/20 sampl es (85%), a distinct IgH-CDR3 PCR product was obtained. Individual PCR products were sequenced after cloning. The nucleotide sequences of 13 4 randomly chosen recombinant vectors were determined demonstrating in 17/20 cases (85%) monoclonal V-H-N-D-H-N-J(H) junctions. Analysis of PCR products by temperature-gradient gel electrophoresis (TGGE) confir med the specificity of the IgH-CDR3 PCR-sequencing results. Moreover, the combination of PCR/TGGE technology allowed the rapid and specific characterization of clonal IgH-CDR3 junctions in B cell proliferations by direct sequencing even in the presence of admired polyclonal B cel ls.