Pitfalls in the diagnosis of diffuse large B-cell lymphoma with flow cytometry

Although flow cytometry (FCM) is not required for the diagnosis of diffuse large B-cell lymphoma (DLBCL) according to World Health Organization criteria, FCM plays an auxiliary role in the work-up of non-Hodgkin lymphomas. The sensitivity of FCM for DLBCL remains variable. Literature reports a detection rate of 63% to 97%. This study aims to identify factors that may influence the sensitivity.

A retrospective data-analysis was performed on 53 patient samples with a diagnosis of DLBCL. A lymphoma screening panel was performed on the same sample as the histopathological diagnosis. Sample quality was compared between cases with and without FCM-detected disease. The immunophenotype of DLBCL in FCM-detected cases was assessed and compared with normal B-cells.

Detection rate was found at 67.9%. Average viability was 57% when disease was detected on FCM, and 28% when it was not. Detection rate in lymphoid samples was 85%, and in non-lymphoid tissue 57%. The immunophenotype of DLBCL was: CD45+ (100%), CD19+ (97%), CD20+ (100%), CD5- (89%), and CD10- (58%). Side scatter was high in most cases, and light chain restriction was not detected in 19%.

This study identifies multiple factors that influence the sensitivity to detect DLBCL cells by flow cytometry in biopsy samples. Non-lymphoid samples of low viability should be considered as limited in diagnostic utility. From an analytical perspective, the gating strategy should be optimized to lower the risk of missing lymphoma cells with a high SSC and the absence of light chain restriction should not exclude the diagnosis of DLBCL in FCM.