T cell activation is a process by which naive T cells differentiate into more specialized roles. This occurs when T cell receptors (TCRs) on the naive cell are stimulated by antigen-presenting cells (APCs) created by the major histocompatibility complex (MHC). Activated T cells work together to protect the body against infectious diseases and other harmful substances. Scientists can study and manipulate these lymphocytes to get a better understanding of how the human body reacts to various pathogens.
Mouse T cell activation occurs in the same way that human T cell activation does within the body. However, researchers can interact with mouse naive T cells to increase activation speeds or alter the type of cell.
Recent tactics have enabled the co-stimulation of T cells with the engagement of CD28, an antigen expressed on most mature T cells. By harnessing this antigen, scientists can reduce the time commitment of T cell activation from 30 hours to 6 hours. This is because co-stimulation of CD28 helps to lower the activation threshold.
With no costimulatory molecules, CD4 T cells require at least 8,000 of their receptors to be engaged before they are activated. In the presence of CD28, only 1,000 to 1,500 TCRs had to be engaged for activation. Whether isolating activated T cells from the mouse or attempting to stimulate and manipulate naive T cells with other antigens, the population may need to be expanded after extraction.
The more cells a researcher can include in their experiment, the higher the chance of accurate results as an outcome. The next step after mouse T cell activation is mouse T cell expansion, which involves culturing target cell populations to increase their quantity. Increasing the population size allows for more tests to be performed and a more accurate representation of how the body would respond in experiments.
While there are reliable methods for large-scale expansion of certain human lymphocytes, such as CD8+ T cells, there are not currently reliable methods for mouse CD8+ T cell expansion. This creates a bottleneck in the research process because scientists must run multiple sorting cycles to isolate the quantity of cells necessary for an experiment. One way the scientific community compensates for the lack of an expansion method is by creating reproducible, generic cell lines that act as the norm for specific cell types.
A cell line is a uniform culture extracted from an organism that is capable of perpetual proliferation. In essence, it’s a representative sample of a greater cell culture that can be expanded continuously so that more cells need not be extracted. When extracted, the sample is still referred to as a primary culture. After the first subculture occurs, the cells will be called a subclone, or cell line. Human cell lines are used to study different bodily systems but can be more difficult to create than mouse cell lines.
A mouse cell line is a culture of murine cells extracted from different organs in a mouse. These are similar to genetic strains, but more consistent and reproducible, allowing researchers to have a basis for studying murine cells. They are created by aggregating several cells and comparing their genetic sequences, then using different programs to filter out mutations or variants that appear to be outliers.
Mouse cell lines provide a biological reference point to compare variables to. It helps researchers to identify which genetic differences may be the cause of certain disorders or mutations. Everything learned from the mouse cell lines can then be used as a reference for human cell lines.
Mouse cancer cell lines hold a cell culture that contains cancer cells. These are engrafted into mice with weakened immune systems so scientists may study how the disease proliferates in them. These mice are efficient hosts due to their decreased immune functions and allow the tumors to grow, producing tumor cell lines and an opportunity to study how cancer affects the mice.
T cells can be found at a very high concentration in the mouse spleen. This organ is a primary target for isolating T cells for cell lines, activation, or expansion. Extracting high volumes of target cells unharmed is an important step when dealing with murine cell research.
Some T cells can be fragile or found in low abundance. To keep a high cell sample viability, it’s important to use a gentle cell separation method. Traditional techniques such as magnetic-based sorting or centrifugation can rupture cell membranes and cause cell death. When cells die, they release cell debris, which can clog up samples and result in exponentially more cell death.
Although these dead cells can be removed with different products, this adds more financial costs and time commitments to the original experiment. Using a cell isolation technique like BACS that does not damage cell health and physiology is a more efficient tactic to maintain sample viability.
Buoyancy activated cell sorting, or BACS, harnesses the buoyant properties of tiny glass bubbles to separate unwanted cells from desired cells in a sample. The bubbles are coated with antibodies that bind to target cells in the solution. Once the cells are linked to the bubbles, they float to the top of the sample where they can be collected with a pipette. Meanwhile, the enriched substance, murine T cells, in this case, remain untouched at the bottom of the sample.
BACS is a cell separation technique developed by Akadeum Life Sciences that was developed to be a fast, easy, and effective method of isolating delicate cells. When creating mouse cell lines out of activated T cells, the primary culture needs to be as healthy as possible. The amount of time a separation method takes can cause cells to die. External forces such as magnets or high-velocity flows can cause cells to die. BACS is a reliable method that will not damage the physiology of desired cells.
BACS also helps to address the issue of T cell expansion. The more cells that can be successfully and precisely isolated, the fewer scientists need to culture them. The 30 to 45-minute workflow can save time, money, and headaches.
Akadeum offers a Mouse T Cell Isolation Kit that uses negative selection to float unwanted cells to the top of a sample. The desired cells can be recovered with up to 96% purity, untouched, and ready for downstream assays. The kit can be used directly in the sample container and requires no additional components.
If you’d like to learn more, check out our page on T cell isolation from mouse spleen or read about how Akadeum’s Mouse T Cell Isolation Kit was used to study vaccine-mediated immunity.