Immunocytometry Assay Services

Make the most of your study samples

Flow cytometry is a powerful technique that allows the analysis of individual cells in a mixed population. By outsourcing your flow cytometry staining experiments to ProImmune, you take advantage of our technical proficiency and save time by passing collection and analysis of data to our experts.

What is Immunophenotyping?

Immunophenotyping by flow cytometry enables expressed proteins on the surface of individual cells to be detected in order to define populations of interest. Cell subset identification may be combined with the analysis of activation status markers, homing markers or exhaustion markers.

ProImmune has experience in designing bespoke immunophenotyping panels, with the capability of detecting up to 8 colours simultaneously. Previous immunophenotyping panels have included:

• T cells including markers to discrimate naive, central memory, effector memory and terminally differentiated effector memory cells: CD3, CD4, CD8, CD45RA, CD62L
• Regulatory T cells: CD3, CD4, CD127, CD25
• B cells: CD19, CD20, CD38, CD27
• Natural killer (NK) cells and NKT cells: CD56, CD3, CD16
• Activation markers: CD69, HLA-DR
• Monocytes and dendritic cells (DCs): CD14, CD16, HLA-DR, CD11b, CD11c, CD209, CD86

We can also include a ‘dump’ channel to maximize color usage, and generally incorporate a live/dead stain. Previous controls include unstimulated samples, isotype controls and Fluorescence Minus One (FMO).

What is Intracellular Cytokine Staining?

Intracellular cytokine staining (ICS) by flow cytometry allows for the analysis of specific T cell responses in defined cell populations. Compared to the ELISpot technique, ICS combines immunophenotyping with the simultaneous detection of cytokines, providing instantly more information about the immune response. In addition, multiple cytokines from the same cell may be detected, facilitating the assessment of polyfunctionality.

ICS relies upon the stimulation of immune cells in the presence of an inhibitor of protein transport to retain the cytokines inside the cell. Cells are first activated with antigen, followed by staining with antibodies specific for particular cell populations, such as CD4 and/or CD8. The cell membranes are then permeabilized prior to staining with fluorochrome-conjugated cytokine-specific antibodies. ICS is commonly used for the detection of human IFN-gamma, IL-2 and TNF-alpha cytokine production in CD4+ and CD8+ T cells.

ICS can be used as an immune monitoring tool to measure the immune response to known antigens, or for epitope discovery, combined with ProImmune’s REVEAL® & ProVE® technology, to identify and/or validate novel T cell epitopes. Read more about ICS

ProImmune is able to analyze either the effect of compounds on immune cells or to assess samples from clinical studies. In this clinical study example, the aim was to compare two cohorts by immunophenotyping PBMCs, identifying classic cell subtypes, and analyzing the cohorts for any differences in the frequency of cell subtypes.