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OMIP-044

28‐color immunophenotyping of the human dendritic cell compartment

Florian Mair, Martin Prlic Cytometry PART A, Volume 93, Issue 4, 402-405 (2018)

PURPOSE: This 28‐color panel has been developed for an extensive phenotyping of antigen‐presenting cells (APCs) in human blood and tissue samples (Table 1). 15 markers associated with changes in dendritic cell (DC) function were selected based on the current literature. Five additional markers were used to pregate on canonical DC subpopulations, namely CD141+ cross‐presenting conventional DC1 (cDC1), CD1c+ conventional DC2 (cDC2), CD141‐ CD1c‐ cDCs as well as CD123+ plasmacytoid DCs (pDC). Furthermore, seven lineage markers were included for parallel enumeration of CD14+ monocytes, B cells, NK cells as well as CD4+ and CD8+ αβ T cells with basic phenotyping of their differentiation status. The panel has been tested on cryopreserved peripheral blood mononuclear cells (PBMCs) and allows further inclusion of DC antigens at the cost of pan‐phenotyping markers.

CELL TYPE: PBMCs and nonlymphoid tissue

MACHINE: BD FACSymphony A5

Marker Target Clone Fluorochrome Purpose
Dead cells Human Live/Dead Fix Blue Viability
CD40 Human 5C3 BUV395 DC phenotype
CD16 Human 3G8 BUV496 DC phenotype, monocyte phenotype
CD56 Human NCAM16.2 BUV563 NK
CD3 Human UCHT1 BUV661 T cell
CD86 Human FUN-1 BUV737 DC phenotype
CD45 Human HI30 BUV805 hematopoitic cell
CX3CR1 Human 2A9-1 Brilliant Violet 421 DC phenotype
CD85k Human ZM3.8 Brilliant Violet 480 DC phenotype
CD45RA Human HI100 Brilliant Violet 570 DC, T cell
CD141 Human 1A4 Brilliant Violet 605 DC subset
CD172a Human SE5A5 Brilliant Violet 650 DC phenotype
CD14 Human MfP9 Brilliant Violet 711 monocyte
CD11b Human ICRF44 Brilliant Violet 750 DC phenotype
CD123 Human 7G3 Brilliant Violet 786 pDC (plasmacytoid DC)
CD26 Human M-A261 FITC DC subset
CD272 Human J168-540 BB630-P2 DC phenotype
CD8 Human RPA-T8 BB660-P2 CD8 T cell
CD32 Human FLI8.26 BB700 DC phenotype
CD38 Human HIT2 BB790-P DC phenotype
ccr7 Human G043H7 PE DC phenotype, T cell phenotype
CD163 Human GHI/61 PE-CF594 DC phenotype
CD80 Human L307.4 PE-Cy5 DC phenotype
CD19 Human SJ25-C1 PE-Cy5.5 B cell
CD4 Human SK3 PE-Cy7 CD4 T cell
CD1c Human F10/21A3 Alexa Fluor 647 DC subset
CD11c Human B-ly6 Alexa Fluor 700 DC lineage marker
HLA-DR Human G46-6 APC-H7 DC lineage marker