Isolation and purification of afferent lymph dendritic cells that drain the skin of cattle
Isolation and purification of afferent lymph dendritic cells that drain the skin of cattle
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DOI：
10.1038/nprot.2006.125
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发表时间：
2006-01-01
期刊：
影响因子：
14.8
通讯作者：
Charleston, Bryan
中科院分区：
文献类型：
--
作者：
Hope, Jayne C.;Howard, Chris J.;Charleston, Bryan
树突状细胞(DC)是诱导免疫反应的中心，也是决定感染挑战结果的关键控制点。通过插入传入淋巴管，可以分离大量的淋巴树突状细胞。首先，通过手术切除正在引流皮肤的淋巴结(大约需要1小时)。在6-8周的时间里，传入淋巴管与传出管道重新吻合，形成可以通过手术插管的更大的假传入淋巴管。外科插管需要2小时；导管的日常维护需要30分钟。分离淋巴细胞需要1小时和额外的60-180分钟来丰富或纯化DC。淋巴可以采集长达1个月，在此期间细胞数量和亚群分布相对恒定。这项技术虽然在技术上要求很高，但有助于研究树突状细胞和其他在稳态和抗原暴露后在淋巴中流动的细胞。
Dendritic cells ( DCs) are central to the induction of immune responses and are a pivotal control point that determines the outcome of infectious challenge. Cannulation of afferent lymphatic vessels allows the isolation of large numbers of lymph DCs. First, lymph nodes that are draining the skin are surgically removed ( takes approximately 1 h). Over a period of 6 - 8 weeks, afferent lymphatic vessels re- anastomose with the efferent duct, forming larger ' pseudoafferent' lymphatic vessels that can be surgically cannulated. Surgical cannulation takes 2 h to perform; daily maintenance of the catheter requires 30 min. Isolation of lymph cells requires 1 h and an additional 60 - 180 min to enrich or purify the DCs. The lymph can be harvested for up to 1 month, with relatively constant cell numbers and subset distribution throughout this period. This technique, although technically demanding, facilitates studies of DCs and other cells that traffic in the lymph in both the steady state and following antigenic exposure.