A 14‐Color, 16‐Antibody Panel for Immunophenotyping Human Innate Lymphoid, Myeloid and T Cells in Small Volumes of Whole Blood and Pediatric Airway Samples

Cytometry PART A, Volume 95, Issue 12, 1231-1235 (2019)
Dawid Swieboda, Yanping Guo, Sophie Sagawe, Ryan S. Thwaites, Simon Nadel, Peter J.M. Openshaw, Fiona J. Culley

Description: This 14‐color, 16‐antibody OMIP was designed for enumeration of leukocyte responses in pediatric samples, where sample volumes and cell numbers can be very low. Leukocytes identified by this panel include all major members of the innate lymphoid cell (ILC) family (ILC1s, ILC2s, and ILC3s), natural killer cells (NK cells), granulocytes (neutrophils and eosinophils), T‐cells (CD4+ and CD8+), mucosal‐associated invariant T cells (MAIT cells) and NKT‐like cells. The protocol was optimized using small volumes of peripheral blood and validated in airway samples obtained from children (< 2 years of age) admitted to a pediatric intensive care unit (PICU). Given this backdrop, this OMIP is widely applicable to clinical research using low volume or paucicellular samples, such as studies of innate and adaptive immune responses in infants and children, with potential clinical application in diagnostics and monitoring of patients by pediatricians.

Species: Human

Tissue: Cord blood, Nasal aspirate, Peripheral blood, Tracheal aspirate

Cell Subset: T cell, Mait, Ilc, Nk, Nkt, Granulocyte, Eosinophil, Neutrophil

Keyword: Pediatric

Machine: BD LSR Fortessa

Panel details

Marker Species Clone Fluorophore Purpose Co-expressions groups Antigen Density
CD14 Human 63D3 Brilliant Violet 510 Lineage   High
CD19 Human HIB19 Brilliant Violet 510 Lineage   Low
CD123 Human 6H6 Brilliant Violet 510 Lineage   Med
CD4 Human RPA-T4 Brilliant Violet 605 CD4+ T cells  T cell High
TCR V alpha 7.2 Human 3C10 Brilliant Violet 785 MAIT cells  MAIT Med
CD45 Human HI30 FITC Leukocyte ILC, T cell, Neutrophil, NK High
CD117 Human A3C6E2 PerCP-Cy5.5 ILC3   ILC Med
CD3 Human OKT3 PE T cells  T cell High
CD161 Human HP-3G10 PE-Dazzle 594 MAIT cells  MAIT, ILC Low
CD56 Human 5.1H11 PE-Cy7 NK/NKT-like cells  NK Low
CD294 Human BM16 Alexa Fluor 647 ILC2/Th2/Tc2 subsets  ILC Med
CD66b Human G10F5 Alexa Fluor 700 Eosinophils  Neutrophil High
Dead cells Human Live/Dead Fix Near IR (775) Viability 
Fc epsilon R1 alpha Human AER-37 (CRA-1) Brilliant Violet 510 Lineage   High
CD8a Human SK1 Brilliant Violet 711 CD8+ T cells  T cell High
CD16 Human 3G8 Brilliant Violet 650 NK cells/neutrophils  NK, Neutrophil Med
CD127 Human A019D5 Brilliant Violet 421 ILCs  ILC Med