CD8<sup>+</sup> Dendritic Cell Isolation Kit ( #130-091-169 )

CD8⁺ Dendritic Cell Isolation Kit

Description

The CD8⁺ Dendritic Cell Isolation Kit is an indirect magnetic labeling system for the isolation of CD8⁺ DCs from single-cell suspensions of spleen and lymph node. CD8⁺ DCs coexpress CD11c, CD205, MHC class II, CD40, CD80, CD86, and are negative for CD4 and CD11b. In spleen and lymph nodes, CD8⁺ DCs are found at moderate levels (23% and 17% of all CD11c⁺ DCs, respectively) and are located in the T cell areas. Skin-draining lymph nodes harbor an additional CD8⁺ DC subset (CD4^-CD8⁺CD11b⁺CD205⁺) which accounts for 33% of all CD11c⁺ DCs in this organ and is thought to represent the mature form of Langerhans cells. The isolation of CD8⁺ DCs is performed in two steps. First, DCs are enriched by depletion of T cells, NK cells, and B cells with a cocktail of biotin-conjugated antibodies against CD90 (Thy1.2), CD49b (DX5), and CD45R (B220) as well as Anti-Biotin MicroBeads. Subsequently, CD8⁺ DCs are positively selected from the enriched DC fraction with CD8a MicroBeads.

Applications

CD8⁺DCs isolated using the CD8⁺ Dendritic Cell Isolation Kit are suitable to analyze antigen uptake, antigen processing, T cell activation or tolerance induction, cross-priming of cytotoxic T cells, and T cell polarization by CD8⁺ DCs. Isolated splenic CD8⁺ DCs have been analyzed for the expression of a signal regulatory protein, which plays a role in the migration of murine Langerhans cells.¹ They additionally have been isolated and adoptively transferred to study their role in the induction of regulatory CD4⁺ T cells.² The CD8⁺ Dendritic Cell Isolation Kit was also used to analyze the expression and regulation of EBV-Induced Gene 3 (EBI3) in mouse DC subsets.³

Columns

For the first magnetic separation (depletion): LD or autoMACS™ Columns; for second magnetic separation (positive selection): MS or autoMACS Columns.

Figure 1

Isolation of CD8⁺ DCs from a mouse spleen cell suspension using the CD8⁺ Dendritic Cell Isolation Kit, an LD and two MS Columns, a MidiMACS™ and a MiniMACS™ Separator.

Spleen cells before separation

Pre-enriched CD8⁺CD11c⁺ DCs

Isolated CD8⁺CD11c⁺ DCs

Figure 2

Working scheme of the CD8⁺ Dendritic Cell Isolation Kit.