>Viki Mosiman >Robert H Lurie Cancer Center >Flow Cytometry Core Facility >Northwestern University Medical School >Chicago, IL >vlm646@nwu.edu > >I am interested in an opinion on the best way to detect Large-cell >lymphoma(B Cell type) in lymph nodes. > >We currently disaggregate nodes then surface stain using >45/19/kappa/lambda. Although >we often have success ,there are cases where clearly the node is a >malignant B-cell lymphoma by morphology in Surgical Pathology but we see >no evidence in Flow. We have tried blocking with fetal calf and mouse >serum. We have tried different gating strategies such as 45ss combined >with 19ss gating or combined with fs. We have tried different staining >combinations -19/5/kappa/lambda-45/19/10/kappa&45/19/10/lambda. We have >tried live dead assays(7AAD) combined with surface staining in case a node >was necrotic. Someone even suggested injecting nodes with PBS to express >cells instead of mincing and grinding lymph nodes(they thought this might >be a more gentle process). > >We don't seem to have the same problem with other malignant lymphomas in >lymph nodes,just with the diagnosis of Large-cell lymphoma ( B-Cell type). >Any suggestions would be greatly appreciated. Thanks for your help! > >Clinical Flow Lab >Northwestern Memorial Hospital > > Viki, Recently we have seen several cases where the malignant population in large cell lymphomas was extremely difficult to demonstrate in flow. One, a lymph node, was easily demonstrated by immunohistology to consist of large numbers of B cells infiltrated with reactive T cells. In flow cytometry after the usual mechanical disaggregation, suspesion in RPMI 1640 + 5% FCS, filtering through 100 micro nylon mesh and washing almost no malignant B cells could be detected. The second was a FNA of a mass in the liver collected into RPMI 1640. This too immunostained positively as large B-cells, but in flow was composed of 99% small to medium T-cells. Cell viability was good in both preparations but there appeared to be substantil very small debris. I'm guessing that these large cells were particularly fragile -- although the appearance of the FNA cells in histology seemed normal. I would be interested in the experience of other with large cell lymphomas. Brent Dorsett Special Pathology Laboratory Lenox Hill Hospital NYC
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