OMIPs Directory

OMIP-067

28‐Color Flow Cytometry Panel to Evaluate Human T‐Cell Phenotype and Function

Cytometry PART A, Volume 97, Issue 10, 1032-1036 (2020)
Phillip A. Swanson II, Robert A. Seder

Description: This 28-color flow cytometry panel was developed to detect frequencies and function of a broad spectrum of T-cell subsets from the blood of infants and adults following antigen-specific stimulation. This panel includes markers to characterize αβ and γδ T-cell subsets, T regulatory cells (Tregs), T follicular helper cells (TfH), and recent thymic emigrants (RTEs). Additionally, antibodies that measure cell surface activation molecules (CD154, CD69, HLA-DR, and CD38) and function through detection of cytokines (IL-8, IL-2, IL-4, IL-5, IL-13, IL-21, IFNγ, and TNF) are also included. This panel was optimized using PMA-stimulated cryopreserved healthy adult and infant PBMCs as well as stimulation with CMV-specific peptides. Currently, this panel is being used to evaluate antigen-specific T-cell responses in adults and infants following immunization with an attenuated malaria vaccine.

Species: Human

Tissue: Peripheral blood

Cell Subset: T cell, Ctl, T helper, T memory, Tcrgd, Th1, Th2, Treg, Recent thymic emigrants (rte), Tfh

Keyword: Activation, Antigen-specific t cell response, Cytokine, Cytomegalovirus (cmv), Infant, Malaria, Pma/ ionomycin

Machine: BD FACSymphony A5

Panel details

Marker Species Clone Fluorophore Purpose Co-expressions groups Antigen Density
TCR V gamma 9 Human IMMU 360 PE-Cy5 Lineage TCRgd Med
Dead cells Human Live/Dead Fix Blue Viability
IL-8 Human G265-8 Brilliant Blue 630-P2 Function Treg Low
PD-1 Human EH12.2H7 Brilliant Blue 660-P2 CO-inhibitory TCRgd, Tfh Low
TCR gamma/delta Human REA591 APC-Vio770 Lineage TCRgd Low
CD185 Human RF8B2 Brilliant Ultraviolet 563 Trafficking TCRgd, Tfh Low
CD69 Human FN50 Brilliant Blue 790-P Activation Tfh, Th1, Th2, TCRgd Med
CD4 Human SK3 Brilliant Ultraviolet 805 Lineage Th1, Th2, Treg Med
CD31 Human WM59 Brilliant Violet 711 Differentiation High
CD3 Human UCHT1 Brilliant Ultraviolet 496 Lineage TCRgd, Th1, Th2, Treg High
CD25 Human BC96 Brilliant Blue 700 Differentiation Treg, Th1, Th2, TCRgd Low
CD27 Human O323 Brilliant Violet 785 Differentiation TCRgd, T memory Med
CCR7 Human G043H7 Alexa Fluor 700 Differentiation T memory, TCRgd Low
CD45RA Human HI100 Brilliant Violet 570 Differentiation T memory, TCRgd Med
CD8a Human RPA-T8 PE Lineage T memory High
CD127 Human A019D5 PE-Cy7 Differentiation Treg Low
CX3CR1 Human 2A9-1 Brilliant Violet 650 Differentiation Th1 Low
CD183 Human G025H7 PE-Dazzle 594 Trafficking Th1 Med
IFN-gamma Human B27 Brilliant Ultraviolet 395 Function Th1, TCRgd High
IL-4 Human MP4-25D2 Brilliant Violet 421 Function Th2, TCRgd Low
IL-5 Human TRFK5 Brilliant Violet 421 Function Th2 Low
IL-13 Human JES10-5A2 Brilliant Violet 421 Function Tfh, Th2 Low
IL-21 Human 3A3-N2.1 Alexa Fluor 647 Function Tfh Low
TCR V delta 1 Human TS8.2 FITC Lineage TCRgd Low
TCR V delta 2 Human 123R3 Vio Green Lineage TCRgd Med
TNF Human Mab11 Brilliant Violet 750 Function TCRgd, Th2 Med
HLA DR Human Tu36 PE-Cy5.5 Activation T memory High
IL-2 Human MQ1-17H12 Brilliant Ultraviolet 737 Function Th1 Med
CD38 Human HIT2 Brilliant Ultraviolet 661 Activation T memory, TCRgd Med
CD40L Human 24-31 Brilliant Violet 605 Co-stimulatory T memory Med