All tags Reagents T cells immunophenotyping

T cells immunophenotyping

T cells play a central role in cell-mediated immunity and their study can help to uncover the mechanisms of many diseases including cancer.

T cells are a type of white blood cell or lymphocyte that recognizes and targets pathogen-infected cells, or cancer cells, for clearance. T cells originate from hematopoietic stem cells in the bone marrow and precursors are shuttled to the thymus for final maturation. T cells are selective, so that they are tolerant to the body’s own cells (self molecules) but remain highly sensitive to non-self pathogens.

Why do we need to study T cells?

Defective T cell function is linked to a number of disease states that can leave a patient severely immunologically compromised. Complete loss of T cell function can be lethal. Studying how T cells recognize and activate immune responses is, therefore, vital for understanding disorders of immune regulation:

Partial T cell disorders render the patient incapable of eliciting a full immune response to an infection.  These disorders may be associated with reduced T cell number or loss of T cell activity. For example, DiGeorge syndrome is a genetic disorder in which the thymus is underdeveloped and recurrent infections are common in patients.

Severe Combined Immunodeficiency (SCID) is an extreme example of immunodeficiency brought on by a number of immune regulation abnormalities, arising from genetic mutations that affect the development and differentiation of T cells and other white blood cells, including B cells.

Autoimmune disease occurs when T cells are unable to distinguish self from non-self and launch an immune response against patients’ own cells. Common disorders such as rheumatoid arthritis are classed as systemic affecting the whole body, whereas organ-specific autoimmune disease may include Crohn’s disease or Celiac disease (gluten intolerance).

There are a number of lymphocyte leukemia types, some of which originate in immature or mature T cells (T cell lymphomas). These cancers can negatively affect red blood cell production and platelets resulting in anemias and leading to blood clotting deficiencies as well as hindering the ability to fight infection. Sometimes T-cell cancer may be virally induced such as in T-cell non-Hodgkin lymphoma, which is linked to HTLV-1 (human T-cell lymphotropic virus 1) infection.

Tumors have developed multiple mechanisms to evade immune cells. However, researchers have now found ways to harness the power of the immune system to combat cancer. These treatments, known as immunotherapies, are showing remarkable responses in patients with advanced stages of the disease. One approach involves using the patient's own immune cells to identify and attack the tumor. T cells are the building blocks of this type of immunotherapy. Chimeric antigen receptors (CARs) have been engineered on the surface of T cells to allow them to recognize specific proteins on tumor cells and attack them. CAR T cells are showing promising results in clinical trials. Another approach is based on the use of monoclonal antibodies, targeting inhibitory immune checkpoint molecules such as PD-L1 to enhance the immune response to tumors. The development of these therapies has been made possible through the study of immune cells, and in particular T cells.

Three subtypes that are commonly studied in T cell research include:

  • Killer T cells (cytotoxic T lymphocytes: CD8+) recognize antigens, released by the infecting agent that becomes bound to the host cell. Killer T cells are capable of identifying and interacting with diverse cell types and pathogens. The T cell receptor binds the foreign antigen triggering the release of cytotoxins into the infected cell leading to cell death.
  • Helper T cells (Th cells: CD4+) facilitate the immune system in fighting infection. Th cells recognize and bind to a pathogenic antigen, leading to the release of soluble factors (cytokines) that signal to the rest of the immune system to launch a response. Th cells are classified into different subsets, including Th1 cells and Th2 cells. Th1 cells bind to phagocytic macrophages and dendritic cells. Th2 cells bind and activate B cells, required for antibody production and, thus, securing life-long immunity against certain bacterial or viral infections.
  • Regulatory T cells (Treg cells: CD4+, CD25+, FoxP3+, CD127+) keep the immune system in check. They ensure that once an immune response has occurred it is shut down. This regulatory mechanism represses immune activity thus preventing an exaggerated immune response. Normally functioning Treg cells are critical for safeguarding against autoimmune disease.

Flow Cytometry: A powerful tool for T cell immunophenotyping

Flow cytometry provides the ability to type immune cells based on their phenotype. The presence of specific cell surface markers, cytokine expression, or phosphorylation of key proteins may be used to immunophenotype specific sub-populations from a heterogeneous starting population. Thus, the unique signature carried by different immune cells can be utilized to isolate cell populations in a similar way to how the immune system targets foreign pathogens: antibody binding.

T lymphocytes may be immunophenotyped on the basis of CD3+ expression and subsequently further sub-divided (e.g. CD8+ Killer T cells and CD4+ Helper T cells). Moreover, T cell phenotypes are flexible and can adapt to different microenvironments; phenotypes may also overlap among multiple T cell populations. Further T cell subset classification may be based on specific cytokine secretion in response to certain stimuli or the phosphorylation of immune signaling proteins, such as STAT proteins. Therefore the ability of the T cell to respond to the environment, such as in a disease state, can be monitored using flow cytometry.

T Cell typeMarkersCellular LocalizationDescription
T cell (all)CD3Cell membraneCell membrane receptor
Killer T cellCD8Cell membraneCell membrane receptor
Killer T cellIFNγ, TNFSecretedCytokines
Killer T cellEOMESNuclearTranscription factor
Helper T cellCD4Cell membraneCell membrane receptor
Helper Th1CXCR3Cell membraneChemokine receptor
Helper Th1IFNγ, IL-2, IL-12, IL-18SecretedCytokines
Helper Th1STAT4, STAT1NuclearTranscription factors
Helper Th2CCR4Cell membraneChemokine receptor
Helper Th2IL-2, IL-4SecretedCytokines
Treg cellCD4, CD25, CD127, CD152Cell membraneCell membrane receptors
Treg cellTGFβ, IL-10, IL-12SecretedCytokines
Treg cellFoxP3, STAT5NuclearTranscription factors


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