Division of Hematology, Mayo Clinic, Rochester, MN.
Division of Hematology, Mayo Clinic, Rochester, MN.
Clin Lymphoma Myeloma Leuk. 2021 Jan;21(1):e48-e51. doi: 10.1016/j.clml.2020.08.003. Epub 2020 Aug 5.
Isolation of malignant plasma cells from bone marrow of patients with monoclonal gammopathies is critical for studies into the disease biology. The plasma cells are typically isolated by positive selection using plasma cell markers such as CD138. Here we have examined the effect of CD138 magnetic bead selection on the expression of other surface phenotypic markers on plasma cells.
Bone marrow aspirates from 16 patients were split and prepared using 2 methods before staining for flow cytometric evaluation. The first method (whole bone marrow) used an ammonium-chloride-potassium lyse of whole bone marrow followed by 2 phosphate buffered saline washes. The second method used CD138-positive magnetic sorting technology (Stem Cell Technology). The cells were run on the FACSCanto flow cytometer after staining for CD38, CD45, CD56, activation markers CD71, CD69, CD154, adhesion markers CD49d, CD49e, CD11a, CD11b, and CD66, B cell markers CD19 and CD20, and for clonality.
There was a substantial loss in the expression of CD71, CD11b, CD11a, CD69, and CD49e on plasma cells following CD138-based sorting. Moreover, in 8 of the 16 cases, there was a nearly complete loss of the CD45-positive subset with a loss of discrimination between CD45-negative and CD45-positive plasma cell subsets in the remaining CD138-sorted preparations.
The change in immunophenotype of the plasma cells on magnetic sorting should be kept in mind when isolating plasma cells using CD138-positive selection for analysis of plasma cells. The technique for characterizing plasma cells should be selected based on the study design to prevent loss of crucial and valuable information.
从单克隆丙种球蛋白病患者的骨髓中分离恶性浆细胞对于研究疾病生物学至关重要。浆细胞通常通过使用浆细胞标志物（如 CD138）进行阳性选择来分离。在这里，我们研究了 CD138 磁珠选择对浆细胞表面其他表型标志物表达的影响。
从 16 名患者的骨髓抽吸物中分离出两份样本，分别采用两种方法进行制备，然后进行流式细胞术评估染色。第一种方法（全骨髓）使用氯化铵-钾裂解全骨髓，然后用 2 次磷酸盐缓冲盐水洗涤。第二种方法使用 CD138 阳性磁分选技术（Stem Cell Technology）。细胞染色后在 FACSCanto 流式细胞仪上运行，用于 CD38、CD45、CD56、激活标志物 CD71、CD69、CD154、粘附标志物 CD49d、CD49e、CD11a、CD11b、CD66、B 细胞标志物 CD19 和 CD20 以及克隆性。
基于 CD138 的分选后，浆细胞上的 CD71、CD11b、CD11a、CD69 和 CD49e 的表达大量丢失。此外，在 16 例病例中的 8 例中，几乎完全丢失了 CD45 阳性亚群，在剩余的 CD138 分选制备物中，CD45 阴性和 CD45 阳性浆细胞亚群之间的区分丧失。
在用 CD138 阳性选择分离浆细胞进行浆细胞分析时，应牢记磁分选后浆细胞免疫表型的变化。应根据研究设计选择用于表征浆细胞的技术，以防止丢失关键和有价值的信息。
