CLONALITY ANALYSIS OF B-CELL LYMPHOMA IN FRESH-FROZEN AND PARAFFIN-EMBEDDED TISSUES - THE EFFECTS OF VARIABLE POLYMERASE CHAIN-REACTION PARAMETERS

To investigate the sensitivity of polymerase chain reaction (PCR)-base d detection of B-cell monoclonality and the effects of several variabl es, we analyzed 119 cases of B-cell lymphomas with proven IgH gene rea rrangements, testing fresh-frozen and formalin-fixed materials in para llel. Using fresh-frozen tissue, 83 cases (70%) were positive with one -step PCR and high-stringency annealing. Two groups of false-negative cases were identified, Group I (16 cases) showing no PCR products, and Group II (20 cases) showing polyclonal smear patterns. Seminested PCR and/or lowered annealing stringency revealed nine additional positive cases in Group I but none in Group II. Amplification with bcl-2/J(H) primers resulted in five more positive cases. The overall positive rat e in the fresh-frozen category was 81% (97 of 119). Parallel analysis was performed on formalin-fixed, paraffin-embedded material from 61 ca ses, and a high concordance rate (89%) was observed. The results indic ated that fresh and formalin-fixed specimens are comparable in this PC R-based assay, and that the two groups of false-negative results can b e accounted for hy different reasons. In addition, we reviewed the cur rent literature, discussed the diagnostic applications of this techniq ue, and listed the core elements of a proposed PCR protocol that shoul d be suitable for most laboratories.