Description
Background information
The isolated PDCs are CD303 (BDCA-2)+, CD304 (BDCA-4/Neuropilin-1)+, CD123+, CD4+, CD45RA+, CD141 (BDCA-3)dim, CD1c (BDCA-1)–, and CD2–. The cells lack expression of lineage markers (CD3, CD14, CD16, CD19, CD20, CD56) and express neither myeloid markers, such as CD13 and CD33, nor Fc receptors, such as CD16, CD64, or FcεRI.
Binding of the antibody to CD304 (BDCA-4/Neuropilin-1) does not have a significant effect on IFN type I production in PDCs. This is in contrast to binding of antibodies to the other PDC-specific antigen, CD303 (BDCA‑2).1, 2
Detailed separation procedure
For the depletion step, all non-PDCs are magnetically labeled by using cocktails containing specific MicroBeads, biotin-conjugated antibodies, and Anti-Biotin MicroBeads. For the subsequent positive selection step, the enriched PDCs are magnetically labeled with CD304 (BDCA-4/Neuropilin-1) Diamond MicroBeads.
Downstream applications
The Diamond Plasmacytoid Dendritic Cell Isolation Kit II was developed for the isolation of PDCs from PBMCs with maximal purity. It is particularly useful if a PDC population with exceptionally high purity is needed as starting material, e.g. for gene expression profiling or protein purification.
Columns
For the first magnetic separation (depletion): LD or autoMACS® Columns. For the second magnetic separation (positive selection): MS, LS, or autoMACS Columns.
Overview
The Diamond Plasmacytoid Dendritic Cell Isolation Kit II yields PDC populations with excellent purities. The kit involves a two-step procedure, consisting of depletion of non-PDCs and positive selection using MicroBeads against the PDC-specific antigen CD304 (BDCA-4/Neuropilin-1). This kit has been developed based on the first-generation kit (# 130-092-402) for even better performance and a shorter protocol.
