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OMIP-055

Characterization of Human Innate Lymphoid Cells from Neonatal and Peripheral Blood

Sabrina Bianca Bennstein, Angela Riccarda Manser, Sandra Weinhold, Nadine Scherenschlich, Markus Uhrberg Cytometry PART A, Volume 95, Issue 4, 427-430 (2019)

PURPOSE: This OMIP panel was designed to characterize the different subsets of human innate lymphoid cells (ILCs) including ILC1, ILC2, and ILC3 within healthy donors' cord blood (CB) (Table 1) in comparison to peripheral blood (PB). As ILCs represent a rare cell population in blood (0.1–0.5% of lymphocytes), the protocol was designed to rigorously exclude the contaminating cells with a complex lineage‐depleting antibody mixture (13 different antibodies) enabling analysis of highly pure ILC subsets by 8‐color staining.

CELL TYPE: Cryopreserved and freshly isolated cord blood and peripheral blood

MACHINE: Beckman Coulter CytoFLEX

Marker Target Clone Fluorochrome Purpose
Dead cells Human Live/Dead Fix Green Viability
CD1a Human HI149 FITC DC
CD14 Human hCD14 FITC Monocyte
CD19 Human HIB19 FITC B cell
CD20 Human 2H7 FITC B cell
CD3 Human UCHT1 FITC T, NKT
TCR alpha/beta Human IP26 FITC CD3-TCRab+
TCR gamma/delta Human B1 FITC exclude CD3-TCR?d
CD123 Human 6H6 FITC DC
CD303 Human 201A FITC pDC (plasmacytoid DC)
Fc epsilon R1 alpha Human AER-37 (CRA-1) FITC master cells, basophils
CD235a Human HI264 FITC erythroid cell and erythroid precursor
CD66b Human G10F5 FITC granulocyte
CD34 Human 581 FITC HSC
CD127 Human R34.34 PE-Cy5 ILC inclusion marker
CD94 Human DX22 PE-Cy7 NK
CD56 Human HCD56 Brilliant Violet 650 NK, ILCs
CD117 Human 104D2 Brilliant Violet 421 ILC3 cell surface receptor
CD294 Human BM16 PE-Dazzle 594 ILC2 cell surface receptor
CD45 Human HI30 APC-Cy7 leucocyte
CD161 Human HP-3G10 Alexa Fluor 700 ILC inclusion marker (especially for tissue)