Simple Panels for the Study of T-Cells
T-cells are part of the adaptive immune system with several subtypes including CD8+ effector T-cells and CD4+ helper T-cells. Main categories can be divided into several sub-categories. What are the critical markers to study T-cells? From the key subsets how can other subsets be defined?
Simplest panel
This panel identifies the main T-cell populations. These markers are expressed at high levels, as a result there is a lot of choice with fluorochrome selection. Very few cells will be double positive for CD4 and CD8.
When building larger panels these markers can be moved to dimmer fluorochromes. CD3 is sometimes removed from panels to make room for more markers as all CD4+ and CD8+ cells are CD3+.
Regulatory T-cells (Treg)
Treg regulate the immune response and are implicated in both cancer and autoimmune disease. In human cells, the most definitive way to identify regulatory T-cells is by utilizing the transcription factor FoxP3. However to access FoxP3 cells must be fixed and permeabilized to access the antigen. Kits and buffer sets, such as True-Nuclear™ Transcription Faction Buffer Set, can simplify experiments and improve results. Once the cells are permeabilized they are not suitable for cell sorting applications. A second Treg panel for sorting is also listed below.
Human Treg analysis panel with FoxP3
Note: Antibodies against FoxP3 are available and work well conjugated to several fluorochromes. One advantage of the Alexa Fluor dyes is their small size. This can be helpful particularly with intranuclear antigens that are part of a complex.
Human Treg sorting panel with CD127
Expression of the IL-7 receptor (CD127) is inversely correlated with FoxP3. CD127 is expressed on the cell surface. Therefore cells do not need to be permeablized and can be sorted. Tregs can be identified as CD4+, CD25+ and CD127-.
There are many additional markers available for the characterization of T-cell subsets. The suggested panels provide a starting point for experiments on entry-level instruments. All reagents should be optimized for best results.
