This application protocol describes the procedure to dissociate mouse spleen, lung, and intestine using the gentleMACS™ Dissociator and specific tissue dissociation kits in order to obtain viable subsets of DCs that can be further isolated using MACS® Cell Separation Technology or flow sorting.
Download kit data sheet
Download kit data sheet
All steps are described in detail in the protocol of the Lamina Propria Dissociation Kit, mouse.
Download kit data sheet
Pan dendritic cells (DCs) are enriched from each of the dissociated tissues with CD11c MicroBeads, UltraPure, mouse. The isolated tissue-derived DCs show specific surface markers that enable the detailed analysis of distinct DC subsets.
Download kit data sheet
Cell surface antigen | Clone | Fluorochrome |
---|---|---|
CD11c | N418 | VioBlue®/APC/PE |
MHC class II | M5/114.15.2 | APC/FITC |
CD11b | REA592 | FITC/APC |
XCR-1 | REA707 | PE |
PDCA-1 | JF05-1C2.4.1 | PE |
Prepare a solution containing phosphate-buffered saline (PBS), pH 7.2, 0.5% bovine serum albumin (BSA), and 2 mM EDTA by diluting MACS® BSA Stock Solution (# 130-091-376) 1:20 with autoMACS® Rinsing Solution (# 130-091-222). Keep buffer cold (2–8 °C).
▲ Note: Always use freshly prepared buffer. Do not use autoMACS Running Buffer or MACSQuant® Running Buffer as they contain a small amount of sodium azide that could affect the results.
Column | Max. number of labeled cells | Max. number of total cells | Separator |
---|---|---|---|
Positive selection | |||
MS | 1×107 | 2×108 | MiniMACS™, OctoMACS™, VarioMACS, SuperMACS II |
LS | 1×108 | 2×109 | MidiMACS™, QuadroMACS™, VarioMACS, SuperMACS II |
XS | 1×109 | 2×1010 | SuperMACS II |
autoMACS | 2×108 | 4×109 | autoMACS Pro |
▲Note: Column adapters are required to insert certain columns into the VarioMACS™ or SuperMACS™ II Separators. For details refer to the respective MACS Separator data sheet.
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