Evaluation of a worldwide EQA scheme for complex clonality analysis of clinical lymphoproliferative cases demonstrates a learning effect

Cleo Keppens,Patricia J T A Groenen,Hongxiang Liu,Elizabeth Hodges,Elisabeth Moreau,Elke Boone,Maria Eugenia Sarasquete,Paul Evans,Dido Lenze,Markus Möbs,Paula Gameiro,Véronique Tack,Elisabeth M C Dequeker,Monika Brüggemann,Ian Carter

doi:10.1007/s00428-021-03046-0

Abstract

Clonality analysis of immunoglobulin (IG) or T-cell receptor (TR) gene rearrangements is routine practice to assist diagnosis of lymphoid malignancies. Participation in external quality assessment (EQA) aids laboratories in identifying systematic shortcomings. The aim of this study was to evaluate laboratories’ improvement in IG/TR analysis and interpretation during five EQA rounds between 2014 and 2018. Each year, participants received a total of five cases for IG and five cases for TR testing. Paper-based cases were included for analysis of the final molecular conclusion that should be interpreted based on the integration of the individual PCR results. Wet cases were distributed for analysis of their routine protocol as well as evaluation of the final molecular conclusion. In total, 94.9% (506/533) of wet tests and 97.9% (829/847) of paper tests were correctly analyzed for IG, and 96.8% (507/524) wet tests and 93.2% (765/821) paper tests were correctly analyzed for TR. Analysis scores significantly improved when laboratories participated to more EQA rounds (p=0.001). Overall performance was significantly lower (p=0.008) for non-EuroClonality laboratories (95% for IG and 93% for TR) compared to EuroClonality laboratories (99% for IG and 97% for TR). The difference was not related to the EQA scheme year, anatomic origin of the sample, or final clinical diagnosis. This evaluation showed that repeated EQA participation helps to reduce performance differences between laboratories (EuroClonality versus non-EuroClonality) and between sample types (paper versus wet). The difficulties in interpreting oligoclonal cases highlighted the need for continued education by meetings and EQA schemes.

Highlights

Clonality testing is widely accepted as a valuable tool in routine diagnosis of lymphoid malignancies [1, 2]
In 2003, the European BIOMED-2 consortium, continued under the name “EuroClonality consortium,” designed a standardized multiplex PCR assay for most IG/T-cell receptor (TR) targets [3], which showed a high rate of clonality detection in common Band T-cell malignancies [4,5,6,7,8,9,10,11,12]
94.9% (506/533) of wet and 97.9% (829/847) of paper tests were correct for IG (MWU, p=0.144), versus 96.8% (507/524) wet and 93.2% (765/821) paper tests for TR (MWU, p=0.686)

Summary

Introduction

Clonality testing is widely accepted as a valuable tool in routine diagnosis of lymphoid malignancies [1, 2]. In 2003, the European BIOMED-2 consortium, continued under the name “EuroClonality consortium,” designed a standardized multiplex PCR assay for most IG/TR targets [3], which showed a high rate of clonality detection in common Band T-cell malignancies [4,5,6,7,8,9,10,11,12]. Reporting of clonality assays is still considered a complex task, because molecular clonality testing reflects immunobiology and comprises the integrated interpretation of multiple multiplex PCR results. These multiplex PCRs use primers of potentially different efficiencies, annealing to highly homologous genes. Interpretation should consider the pathology and the clinical question as the presence of a small clone in a reactive lesion has another implication than its presence in a full-blown malignancy

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