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Stephen P Robinson, MD, PhD, and Andrew Stagg, PhD, have brought together a wide range of time-proven methods for studying these dendritic cells. Many of these readily reproducible techniques deal with the problem of obtaining sufficient dendritic cells for analysis, whether by isolation from a wide vareity of tissues, or from various progenitor cell populations. Other methods describe in step-by-step fashion the techniques commonly used for analyzing aspects of dendritic cells, ranging from cell migration to antigen uptake and T cell stimulation. In addition, a few techniques explore the practical challenges involved in using dendritic cells in a clinical setting to develop novel immunotherapeueutics.
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M E T H O D S I N M O L E C U L A R M E D I C I N E TM Dendritic Cell Protocols Edited by Stephen P. Robinson, MD, PhD Andrew J. Stagg, PhD Humana Press DC from Mouse Lymph Nodes 3 1 Isolation of Dendritic Cells from Mouse Lymph Nodes Dmitry Gabrilovich 1. Introduction Lymph nodes are the primary sites of T-cell stimulation by dendritic cells (DC). After contact with antigens, DCs migrate to draining lymph nodes from the skin and other tissues (1–3). Investigation of the morphology and function of lymph node DCs may provide important information about the role of these cells in normal and pathological conditions. Therefore, lymph nodes are popular sites for the isolation of dendritic cells. Dendritic cells isolated from lymph nodes represent “interdigitating” DCs that are localized in T-dependent regions of lymph nodes. DCs represent about 1% of the total population of lymph node cells. Therefore, in order to perform almost any functional tests, the DC fraction should be enriched. The most practical way to enrich the DC fraction is to use a density gradient. Several gradients—metrizamide (4), Nycode