Description

About the Cynomolgus Monkey (or Cynomolgus Macaque)

The cynomolgus macaque is the most commonly utilized non-human primate in biomedical research.  According to some researchers, these monkeys are ideal models due to the 90-93% genetic similarity to and recent evolutionary divergence from humans.  They are employed in numerous research areas, such as immunology, neuroscience, oncology, diabetes, and pharmacology due to their genetics and physiology.

Cynomolgus Macaque Monkey CD4 T Cells

Cynomolgus CD4 T cells make up approximately 25-47% of PBMCs in a typical adult monkey. These cells carry the CD4 protein marker and the T cell receptor (TCR), which recognizes foreign antigens presented by major histocompatibility complex class II molecules on antigen presenting cells to activate them.

CD4 T cells develop in the thymus to become mature T cells and exit the thymus to reside and encounter foreign pathogens in lymphoid tissues.

When activated, CD4 T cells become T helper subsets to secrete cytokines to boost the immune response. Depending on the helper subset and cytokine released, the humoral (antibody-based) or cell-mediated immune response is elaborated.

CD4 T Cell Application Summary

Purified cynomolgus CD4 T cells are a good source of cells to study the biology of CD4 T cells and their role in the immune system. They can be used for a variety of stimulation-dependent functional experiments to assess cytokine production and proliferation.

With the current trend toward development of immuno-modulatory drugs, CD4 T cells are an ideal source of cells to test cytokine response in the presence of therapeutic molecules.  More importantly, cynomolgus T cells can be used to assess toxicity and safety characteristics of these biologics in the presence of CD4 T cell activation before first in-human trials.  These cells are commonly employed in pre-clinical settings to ensure biologics are not eliciting unwanted functions, specifically from CD4 T cells.

The negative isolation of CD4 T cells leaves them untouched without any antibody binding to cell surface markers that may influence function. This method leaves all cell surface proteins eligible to be bound to antibodies or other molecules for functional or population characterization studies.

In contrast, positive isolation of CD4 T cells may lead to internalization of the marker that was used to isolate the cells.  In most cases, these markers are only used for identification purposes and may not have any effect on function, but it will depend on the organism and function.  Therefore, cells isolated using this method may also be employed for functional or population characterization studies with the knowledge that the isolation marker may be internalized and not present.

CD4 T Cell Purification

Collection of samples

Cynomolgus PBMCs were sourced from a responsible third party vendor that operate according to local regulations and laws.

Isolation of Cyno CD4 T cells

To enrich for cyno CD4 T cells by the negative isolation method, PBMCs were incubated with antibodies against CD8 T cells, B cells, NK cells, dendritic cells, monocytes, granulocytes, and erythrocytes, and subsequently subjected to a magnet. Cells labeled with the antibodies bound to the magnet through the test tube wall, while unlabeled cells, the CD4 T cells, were decanted into a fresh tube to obtain the enriched population.

For the positive selection method, PBMCs were incubated in a test tube with antibodies against a CD4 T cell-specific marker and subsequently subjected to a magnet, similar to negative isolation.  However, the cells that were decanted into the fresh test tube were the non-CD4 T cells, which can be used for other purposes, while the labeled CD4 T cells were left bound to the magnet through the test tube wall.  The test tube was then removed from the magnet to release the purified CD4 T cells directly into the test tube.

Contact us for more information about purchasing our magnetically isolated CD4 T cells.

Purity

A small aliquot of Cyno CD4 T cells was tested for post-sort purity by flow cytometry analysis. Purity of CD4 T cells, as defined by CD4 expression, was > 97% (Figure 1).

Figure 1. Purity of Cyno CD4 T cells after negative magnetic isolation from PBMCs.  In this representation, cynomolgus PBMCs were incubated with antibodies against CD8 T cells, B cells, NK cells, dendritic cells, monocytes, granulocytes, and erythrocytes and subjected to negative magnetic isolation to obtain untouched CD4 T cells.  A small aliquot was taken after selection to evaluate post-sort purity (right).  Percent of CD4+ cells in PBMCs is also shown (left).

Cryopreservation and storage

Purified CD4 T cells were cryopreserved carefully using iQ Biosciences’ cryopreservation protocol that ensures high viability (> 90%) after thawing.

Cells should be stored at < -120 °C once they are received, such as within a liquid nitrogen tank (vapor phase).