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. 2016 Apr 8;5(4):e68. doi: 10.1038/cti.2016.5

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Copyright © 2016 Australasian Society for Immunology Inc.

This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International License. The images or other third party material in this article are included in the article's Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by-nc-nd/4.0/

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Validation of SPADE clustering to identify discreet DC subpopulations in the combined analysis of purified and PE-conjugated test mAbs. Manually gated bivariate plots for submitted mAbs to well-described monocyte- and DC-specific molecules were compared with manually annotated SPADE trees generated using the DC backbone. Each SPADE tree was interrogated for binding of well-described monocyte or DC markers. (a) CD300a (clone MEM260) was used as a pan myeloid marker, (b) ILT7 (clone 17G10.2) identified pDCs co-expressing CD304 (clone 7G3), (c) CD1c (clone L161) identified DCs co-expressing BDCA-1 (clone AD5-8E7) and (d) CLEC9A (clone 9A11) identified DCs co-expressing CD141⁺ (clone AD54H12). CM,CD14⁺⁺ CD16⁻ classical monocytes; DCs; CD1c⁺ DCs; CD141⁺ DCs and CD16⁺ DCs are gated as the CD11c⁺ mDC subsets, mDC,CD11c⁺CD304⁻ myeloid, NCM,CD14⁺ CD16⁺⁺ non-classical monocytes, pDC,CD304⁺ plasmacytoid DCs.