Flow cytometric analysis of canine lymphoma samples stored in a preservative medium for up to 7 days

Background Fresh samples are required for flow cytometry (FC). A recent study documented the stability of lymphoid markers in peripheral blood samples of healthy dogs stored in a preservative medium up to 7 days. The effects of preservation on canine lymphoma cells in specific mediums are unknown. Objective To assess the quality for FC of lymphoma samples preserved in Transfix® (Cytomark, Buckingham, UK) for up to 7 days from sampling. Methods Lymph node (LN) aspirates from dogs with lymphoma were collected into RPMI tubes and Transfix-tubes. Mean fluorescence intensity (MFI) of unstained cells and of CD45-, CD5-, CD21-, CD4- and CD8-stained cells was recorded, together with the percentage of positive cells. Data from fresh samples collected in RPMI and preservative tubes were compared. Samples stored in the Transfix-tubes were processed and analyzed daily up to 7 days from sampling, and data were compared with a Friedman test. Results It was possible to diagnose lymphoma from all samples at any time, and percentages of positive cells remained constant. Morphological parameters of Transfix-samples were strongly affected. MFI of unstained cells was higher in Transfix- than in RPMI-samples, and increased with time. MFI of antibodies was lower in Transfix- than in RPMI-samples, and decreased with time. Conclusion Transfix Cytomark® is suitable for diagnostic purposes in canine lymphoma samples up to 7 days. However, fresh RPMI samples should be preferred, as they provide better morphological assessment. Also, low degree of antigen expression may be undetectable in Transfix-samples, possibly affecting diagnosis.