Isolation of functionally active murine follicular dendritic cells
Isolation of functionally active murine follicular dendritic cells
复制标题
DOI：
10.1016/j.jim.2006.03.018
复制
发表时间：
2006-06-30
影响因子：
2.2
通讯作者：
Tew, John G.
中科院分区：
文献类型：
--
作者：
Sukumar, Selvakumar;Szakal, Andras K.;Tew, John G.
滤泡树突状细胞（FDCs）的生物化学，遗传学和免疫学研究一直受到阻碍，难以获得足够数量的纯化细胞的功能状态。为了解决这一障碍，我们通过照射小鼠来破坏大多数淋巴细胞，切除淋巴结，并用酶混合物轻轻消化淋巴结以形成单细胞悬液来富集FDC。用特异性单克隆抗体FDC-M1和磁性细胞分离技术选择悬浮液中的FDCs。我们能够以约90%的纯度获得每只小鼠近一百万个活的淋巴结FDC。当在光镜和透射电镜下检查时，细胞学特征是FDC的特征。此外，细胞能够捕获和保留免疫复合物，并且对重要的表型标记物呈阳性，包括FDC-M1、CD 21/35、CD 32、CD 40和CD 54。此外，纯化的FDC表现出经典的FDC辅助活性，包括：共刺激B细胞增殖，增强由有丝分裂原或抗原诱导的抗体应答，维持B细胞存活数周，以及保护B淋巴细胞免受抗FAS诱导的凋亡的能力。简而言之，这种方法的组合使得有可能获得大量高度富集的功能性鼠FDC。(c)2006 Elsevier B. V.保留所有权利。
Biochemical, genetic, and immunological studies of follicular dendritic cells (FDCs) have been hampered by difficulty in obtaining adequate numbers of purified cells in a functional state. To address this obstacle, we enriched FDCs by irradiating mice to destroy most lymphocytes, excised the lymph nodes, and gently digested the nodes with an enzyme cocktail to form single cell suspensions. The FDCs in suspension were selected using the specific mAb FDC-M1 with magnetic cell separation technology. We were able to get nearly a million viable lymph node FDCs per mouse at about 90% purity. When examined under light and transmission electron microscopy, the cytological features were characteristic of FDCs. Furthermore, the cells were able to trap and retain immune complexes and were positive for important phenotypic markers including FDC-M1, CD21/35, CD32, CD40, and CD54. Moreover, the purified FDCs exhibited classical FDC accessory activities including: the ability to co-stimulate B cell proliferation, augment antibody responses induced by mitogens or antigens, maintain B cell viability for weeks, and protect B lymphocytes from anti-FAS induced apoptosis. In short, this combination of methods made it possible to obtain a substantial number of highly enriched functional murine FDCs. (c) 2006 Elsevier B.V. All rights reserved.