CD1c (BDCA-1)+ Dendritic Cell Isolation Kit, human

CD1c (BDCA-1)
+
Dendritic Cell Isolation Kit
, human

The CD1c (BDCA-1)
+
Dendritic Cell Isolation Kit has been developed for the isolation of CD1c (BDCA-1)
+
myeloid dendritic cells (MDCs; type-1 MDCs or MDC1s) from PBMCs.

Background information

The CD1c (BDCA-1) antigen is specifically expressed on dendritic cells, which are CD11c
high
CD123
low
and represent the major subset of MDCs in human blood (about 0.6 % of all PBMCs).
1–7
CD1c (BDCA-1)
+
MDCs show a monocytoid morphology and express myeloid markers such as CD13 and CD33 as well as Fc receptors such as CD32, CD64, and FceRI. Furthermore, they were determined to be CD4
+
, Lin (CD3, CD16, CD19, CD20, CD56)
–
, CD2
+
, CD45RO
+
, CD141 (BDCA-3)
low
, CD303 (BDCA-2)
–
, and CD304 (BDCA-4/Neuropilin-1)
–
.
1
A minor proportion of CD1c (BDCA-1)
+
MDCs expresses CD14 and CD11b. CD1c (BDCA-1) is also found on CD1a
+
dendritic cells generated
ex vivo
from monocytes or hematopoietic precursor cells. In blood, apart from MDCs, a subset of B cells also expresses CD1c (BDCA-1). Therefore, the CD1c (BDCA-1)
+
Dendritic Cell Isolation Kit includes CD19 MicroBeads for depletion of B cells prior to the enrichment of CD1c (BDCA-1)
+
MDCs. In order to discriminate the CD1c (BDCA-1)
+
from CD1c (BDCA-1)
–
CD141 (BDCA-3)
++
MDCs, the former have been designated type-1 MDCs (MDC1s).
The CD1c antigen is a member of the CD1 family of proteins that are structurally related to MHC class I proteins and mediate the presentation of non-peptide antigens towards T cells.
14

Detailed separation procedure

First, B cells are depleted with CD19 MicroBeads. Then, MDC1s are enriched by magnetic labeling with biotin-conjugated CD1c antibodies and Anti-Biotin MicroBeads.

Downstream applications

Isolated CD1c (BDCA-1)
+
MDCs were used to examine the expression of Toll-like receptors
4, 5, 12
, chemokine receptors
3, 5, 8
, or new antigens, e.g., DCAL-16 and EMR27. Further, they were isolated for studies on dendritic cell activation
4
, migration
3
, cytokine production
4, 5, 10
, and T cell polarization
4
, particularly in comparison with monocyte-derived dendritic cells
9
. The CD1c (BDCA-1)
+
Dendritic Cell Isolation Kit was also used for isolation of MDCs from mice engrafted with human CD34
+
hematopoietic progenitor cells
11
, for gene clustering of dendritic cell subsets in blood and tonsils
15
, for phenotypical analysis of MDCs in PBMCs and synovial fluid of psoriatic arthritis and rheumatoid arthritis patients
16
, or as an alternative to CD1a MicroBeads for the isolation of epidermal Langerhans cells
13
.

Columns

For the first magnetic separation (depletion): LD or autoMACS
®
Columns. For the second magnetic separation (positive selection): MS, LS, or autoMACS Columns.

  • Figure 1

    Isolation of CD1c (BDCA-1)
    +
    myeloid dendritic cells isolated from PBMCs by using the CD1c (BDCA-1)
    +
    Dendritic Cell Isolation Kit, a VarioMACS™ Separator, LD and MS Columns. Cell samples of each separation step were stained for CD19 and indirectly for CD1c (BDCA-1) using Anti-Biotin antibodies.
    PBMCs before separation
    Enriched CD1c (BDCA-1)
    +
    dendritic cells after B cell depletion
    Isolated CD1c (BDCA-1)+ myeloid dendritic cells

  • Figure 2

    View details

    Figure 2

    Working scheme for the isolation of CD1c (BDCA-1)
    +
    myeloid dendritic cells from peripheral blood using the CD1c (BDCA-1)
    +
    Dendritic Cell Isolation Kit.
    Working scheme for the isolation of CD1c (BDCA-1)
    +
    myeloid dendritic cells from peripheral blood using the CD1c (BDCA-1)
    +
    Dendritic Cell Isolation Kit.

  • Selected references

    1. Dzionek, A. et al. (2000) BDCA-2, BDCA-3, BDCA-4: Three markers for distinct subsets of dendritic cells in human peripheral blood. J. Immunol. 165: 6037-6046
    2. Grabbe, S. et al. (2000) Dendritic cells: multi-lineal and multi-functional. Immunol. Today 21: 431-433
    3. Penna, G. et al. (2001) Cutting Edge: Selective usage of chemokine receptors by plasmacytoid dendritic cells.. J. Immunol. 167: 1862-1866
    4. Krug et al. (2001) Toll-like receptor expression reveals CpG DNA as a unique microbial stimulus for plasmacytoid dendritic cells which synergizes with CD40 ligand to induce high amounts of IL-12. Eur. J. Immunol. 31: 3026-3037
    5. Jarrossay et al. (2001) Specialization and complementarity in microbial molecule recognition by human myeloid and plasmacytoid dendritic cells. Eur. J. Immunol. 31: 3388-3393
    6. Ryan, E. J. et al. (2002) Dendritic cell-associated lectin-1: a novel dendritic cell-associated, C-type lectin-like molecule enhances T cell secretion of IL-4. J. Immunol. 169: 5638-5648
    7. Kwakkenbos, M. J. et al. (2002) The human EGF-TM7 family member EMR2 is a heterodimeric receptor expressed on myeloid cells. J. Leukoc. Biol. 71(5): 854-862
    8. Penna, G. et al. (2002) Cutting edge: differential chemokine production by myeloid and plasmacytoid dendritic cells. J. Immunol. 169: 6673-6676
    9. Jefford, M. et al. (2003)
      Functional comparison of DCs generated
      in vivo
      with Flt3 ligand or
      in vitro
      from blood monocytes: differential regulation of function by specific classes of physiologic stimuli.
      Blood 102: 1753-1763
    10. Steinschulte, C. et al. (2003) Cutting edge: sanglifehrin A, a novel cyclophilin-binding immunosuppressant blocks bioactive IL-12 production by human dendritic cells. J. Immunol. 171: 542-546
    11. Palucka, A. K. et al. (2003)
      Human dendritic cell subsets in NOD/SCID mice engrafted with CD34
      +
      hematopoietic progenitors.
      Blood 102: 3302-3310
    12. Matsumoto, M. et al. (2003) Subcellular localization of Toll-like receptor 3 in human dendritic cells. J. Immunol. 171: 3154-3162
    13. Peiser, M. et al. (2003) CD1a and CD1c cell sorting yields a homogeneous population of immature human Langerhans cells. J. Immunol. Methods 279: 41-53
    14. Brigl, M. and Brenner, M. B. (2004) CD1: antigen presentation and T cell function. Annu. Rev. Immunol. 22: 817-890
    15. Lindstedt, M. et al. (2005)
      Gene family clustering identifies functionally associated subsets of human
      in vivo
      blood and tonsillar dendritic cells.
      J. Immunol. 175: 4839-4846
    16. Jongbloed, S. L. et al. (2006) Enumeration and phenotypical analysis of distinct dendritic cell subsets in psoriatic arthritis and rheumatoid arthritis. Arthritis Res. Ther. 8: R15

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