Vos Jeffrey A, Simurdak Jerry H, Davis Brad J, Myers Jerome B, Brissette Mark D
Department of Pathology, Madigan Army Medical Center, Tacoma, Washington, USA.
Cytometry B Clin Cytom. 2003 Mar;52(1):20-31. doi: 10.1002/cyto.b.10002.
Many approaches to obtaining single cells from tissue for flow cytometric immunophenotyping are used; however, these methods result in tissue that is too disrupted for subsequent histologic examination. We introduce a new technique for cell dissociation of hematopoietic malignancies that preserves tissue for histology. This is especially important with small specimens for which this type of correlation is critical.
Fresh tissue from lymph node, gastrointestinal (GI) tract, skin, and other soft tissue biopsies, in addition to cores of inaspirable bone marrows, were briefly vortexed until the RPMI cell culture medium became cloudy. Larger specimens such as lymph nodes were sectioned before disaggregating, whereas smaller ones were vortexed in toto. Resultant flow cytometric analyses were compared with the histology and, in some cases, the immunohistochemistry (IHC) to determine whether the data were concordant. Cell suspensions of 104 specimens-composed of 48 lymph nodes, 19 bone marrow cores (BMCs), 11 GI biopsies, 11 skin/soft tissue biopsies, and 15 miscellaneous specimens-were prepared via vortex disaggregation.
Flow cytometric analysis of 96 specimens (92.3%) showed adequacy of material and diagnostic correlation with the histology and IHC. Of the eight cases (7.7%) that were discordant, seven were attributable to significant specimen fibrosis or necrosis. With respect to tissue type, this method produced diagnostic cell suspensions for most lymph nodes (95.8%), GI biopsies (90.9%), and BMCs (89.5%); however, it was less useful for skin/soft tissue samples (81.8%).
Disaggregation of tissue for flow cytometric analysis by vortexing appears to provide adequate and representative cellular material. This technique is ideal for inaspirable bone marrows and small biopsies where tissue preservation for histology is paramount.
目前有多种从组织中获取单个细胞用于流式细胞术免疫表型分析的方法；然而，这些方法会导致组织过度破碎，无法进行后续的组织学检查。我们引入了一种用于造血系统恶性肿瘤细胞解离的新技术，该技术能保留组织用于组织学检查。对于此类相关性至关重要的小标本而言，这一点尤为重要。
除了无法抽吸的骨髓芯外，来自淋巴结、胃肠道（GI）、皮肤及其他软组织活检的新鲜组织，经短暂涡旋直至RPMI细胞培养基变浑浊。较大的标本如淋巴结在解离前进行切片，而较小的标本则整体涡旋。将所得的流式细胞术分析结果与组织学结果进行比较，在某些情况下还与免疫组织化学（IHC）结果进行比较，以确定数据是否一致。通过涡旋解离制备了104个标本的细胞悬液，其中包括48个淋巴结、19个骨髓芯（BMC）、11个GI活检标本、11个皮肤/软组织活检标本和15个其他标本。
96个标本（92.3%）的流式细胞术分析显示材料充足且与组织学和IHC诊断相关。在8个不一致的病例（7.7%）中，7个归因于标本显著纤维化或坏死。就组织类型而言），该方法为大多数淋巴结（95.8%）、GI活检标本（90.9%）和BMC（89.5%）产生了诊断性细胞悬液；然而，对皮肤/软组织样本的效果较差（81.8%）。
通过涡旋进行组织解离以用于流式细胞术分析似乎能提供充足且具有代表性的细胞材料。该技术对于无法抽吸的骨髓和小活检标本非常理想，因为在这些情况下，保留组织用于组织学检查至关重要。
