Improved detection of diffuse large B‐cell lymphoma by flow cytometric immunophenotyping—Effect of tissue disaggregation method

Abstract

Background: Flow cytometric immunophenotyping (FCI) is recognized as a rapid, sensitive, and accurate method for diagnosis of B-cell lymphomas. We observed that FCI failed to identify the clonal B-cell population in several cases of large B-cell lymphoma (DLBCL) when tissue samples were prepared by a commercially available mechanical tissue disaggregation method. We tested a manual tissue disaggregation method and compared it with the mechanical method.Methods: FCI findings from 51 cases of DLBCL processed with the mechanical tissue disaggregation method, 27 cases processed using the manual method, and 15 cases processed using a combination of both methods were compared. The histological and immunohistochemical findings in each case were reviewed.Results

Evaluation of primary mediastinal large B cell lymphoma by flow cytometry

“…Furthermore, the sensitivity of the B-NHL assay reported in the study to detect the neoplastic population of PMLBCL (75%) is in line with prior rates of DLBCL, NOS detection by flow cytometry (27). Interestingly, while both assays for B-NHL and CHL could detect the neoplastic cells in most cases, the CHL tube was superior.…”

“…While it is difficult to say with certainty why the B-NHL failed to identify the neoplastic population of PMLBCL, the fact that the neoplastic population could be identified in 4 of 5 cases implies that preanalytical variables (e.g., loss of neoplastic cells in processing) are likely not the explanation for this finding. Furthermore, the sensitivity of the B-NHL assay reported in the study to detect the neoplastic population of PMLBCL (75%) is in line with prior rates of DLBCL, NOS detection by flow cytometry (27). Importantly, the neoplastic populations from these cases initially interpreted as falsely negative using the B-NHL tube were all present at less than 1% of the total cells evaluated (range, 0.12-0.81; mean, 0.51% for all cells) and were light chain negative, likely playing a role in the initial false negative result.…”

An atlas of B-cell clonal distribution in the human body

“…As different B cell subsets differ in their antibody RNA transcript levels, are not fully defined in human tissues and vary in their ease of recovery in single cell suspensions from tissues, we extracted DNA from whole tissue samples. 15 The analysis of DNA permitted efficient (one template per cell), large-scale, and agnostic sampling of all B cells. Antibody heavy chain gene rearrangements were amplified and sequenced (see Methods and Supplementary Tables 1 and 2 ).…”

The current role of clinical flow cytometry in the evaluation of mature B‐cell neoplasms

“…In addition, flow cytometric studies are less often successful in the evaluation of large B-cell lymphomas, possibly due to increased cell turnover, tumor necrosis and poor cell preservation, and might be improved by rapid processing and gentle disaggregation (50,51). Usually large B-cell lymphoma can be readily distinguished from a reactive process by morphology, and the immunophenotype has a less prominent role to play in sub-classification.…”

“…Usually large B-cell lymphoma can be readily distinguished from a reactive process by morphology, and the immunophenotype has a less prominent role to play in sub-classification. In addition, flow cytometric studies are less often successful in the evaluation of large B-cell lymphomas, possibly due to increased cell turnover, tumor necrosis and poor cell preservation, and might be improved by rapid processing and gentle disaggregation (50,51).…”