The Agilent Advantage – Measure Immune Cell Activation in Minutes
Cellular energy metabolism is now understood to influence many immune cell
pathways as both an indicator and controller of immune cell function and
lineage commitment. This concept is clearly illustrated in the activation
of naive T cells to stimulate proliferation and macrophages to stimulate
Expand these sections below to learn more about accelerating your immune cell research with XF technology.
Immune Cell Activation Application Notes & Technical Information
T CELL ACTIVATION
Real-time analysis of in situ activated T cells
Measure and Discriminate T cell
Activation in Real-Time
Based on recent discoveries, glycolytic pathway dependency for energy production is a robust measure of T cell activation. Conventionally, progression of T cell activation is measured in terms of changes in cell size/morphology, interleukin/interferon expression and cell surface markers, methods which can be time and labor intensive, often involve end-point (not kinetic) data, and a time scale of hours to days. The Seahorse XFp analyzer can detect T cell activation response within several minutes of stimulation with CD3/CD28 immunogenic beads by measuring glycolytic extracellular acidification rate (ECAR) in real time providing an early window of functional information to discriminate activation responses.
Real-time analysis of in situ activated macrophages
Measure and Discriminate Macrophage Activation in Real-Time
Based on recent discoveries, glycolytic pathway dependency for energy production is a robust measure of macrophage activation.
Conventionally, progression of macrophage activation is measured in terms of changes in cytokine expression and other end-point (not kinetic) data. The Seahorse XFp analyzer can detect macrophage activation response, stimulated with LPS using our integrated injection ports, by measuring the proton efflux rate (PER) in real time providing an early window of functional information to discriminate activation responses.
Real-time analysis of in situ activated neutrophil
Direct Detection of Neutrophil
Oxidative Burst in Real-Time
Neutrophils are the initial immune defense recruited to site of infection and injury, they have several mechanisms to eliminate infections among them production of reactive oxygen species (ROS), phagocytosis and release extracellular traps (NET). Conventionally, ROS production is measured by fluorescence or luminescence detection, methods that are indirect, non-quantitative, not specific, and prone to artifacts.
Agilent Seahorse XF technology enables real time detection and quantification of neutrophil activation by simultaneously measuring the oxygen consumption rate (OCR), oxygen consumed when ROS are generated, and proton efflux rate (PER; a measure of glycolysis) required for ROS production. Bioenergetic response as measured by XF technology is required for neutrophils to meet their rapidly changing cellular energy and metabolite demands.
Simultaneous measurements of OCR and PER are also not possible with other methods. Within the application note, the utility of the XF neutrophil activation assay can be expanded to enable studies of innate immune responses to modulators, drug treatments and micro environment cues.
Real-time discrimination of immune cell activation responses.
Introduce up to 4 modulators per well with programmable drug injection ports to elucidate immune cell response to activation kinetics.
Explore the metabolism driving immune cell response through assays providing metabolic phenotype, bioenergetic pathway, and fuel/substrate preference.
Adapted from Hongbo Chi, Nat Rev Immunol.;
12(5): 325-338, 2012
The Cell Lineage and Immunometabolism Link
Advances in the field of immunology show cellular metabolism as a driver and regulator of immune cell function and differentiation.
T cell and macrophage activation are correlated with metabolic shifts toward glycolysis, activating energy promoting pathways required for high rates of proliferation or signaling.
Modulate immune cell activation
with the use of programmable drug injection ports
Agilent Seahorse XF sensor cartridges incorporate integrated injection ports that sequentially deliver up to 3 compounds, in addition to immunogenic activators, allowing dose response, agonist or antagonist response, or pathway perturbation analysis of each sample.
Dive deeper with standardized cell metabolism assays and powerful software
In addition to T cell and macrophage activation, Seahorse metabolic assay kits are available for measuring mitochondrial respiration, glycolysis, and fatty acid oxidation in cells. Seahorse XF kits and reagents are pre-calibrated, pre-tested, and are coupled with powerful report generator software, providing functional metabolic data from live immune cells.
Rapidly determines the phenotype/fingerprint of cells and quantify ATP production rates