Frequency, interobserver reproducibility and clinical significance of equivocal peaks in PCR clonality testing using Euroclonality/BIOMED-2 primers.

Abstract

AIMS: PCR studies for lymphoid clonality are now widely employed, especially

using Euroclonality/BIOMED-2 primers. Criteria for interpretation as a clonal

result, however, have proven controversial. This study examines the frequency and

clinical significance of equivocal amplification patterns and measures the

interobserver reproducibility of clonality interpretations. METHODS: At our

institution, results of each primer set are first classified as clonal,

non-clonal or abnormal (equivocal peak on polyclonal background). Final results

for all primer sets are then collectively reported as positive (>/=1 clonal

result), negative (non-clonal results) or indeterminate (>/=1 abnormal result)

for a clonal population. Results of 274 consecutive clonality cases were

reviewed, and the interobserver reproducibility of individual primer set

reactions and final results was determined in a subset of 30 cases. RESULTS:

44/161 (27%) B-cell and 50/163 (31%) T-cell cases contained at least one abnormal

peak. Of these, 29 (64%) and 31 (62%), respectively, showed clonal results in

another primer set. Interobserver reproducibility was excellent for most primer

sets and for final interpretations, but only fair to good for IGK V-J and TCRB

D-J1+2 primer sets. A definitive diagnosis of lymphoma was rendered in 93%, 20%

and 6% of B-cell cases and 90%, 42%, and 14% of T-cell cases positive,

indeterminate or negative for a clonal population, respectively. CONCLUSIONS:

Using a subjective approach, abnormal (equivocal) peaks are frequently observed

in routine practice. However, most cases with abnormal peaks contain clonal

rearrangements in other primer sets, facilitating overall interpretation of final

results with excellent interobserver reproducibility.