Peripheral blood mononuclear cells (PBMCs) are a collection of immune cells that circulate in the blood. Scientists freeze these cells through a process called cryopreservation, which allows them to be stored for extended periods for future research. This use of frozen PBMCs gives researchers flexibility, enabling them to conduct experiments when it is most convenient and to ensure consistency in studies that span long durations.
Understanding Peripheral Blood Mononuclear Cells
Peripheral blood is the term for blood that circulates throughout the body, as opposed to blood stored within the bone marrow. A “mononuclear cell” refers to any blood cell that has a single, round nucleus. The main types of PBMCs are lymphocytes and monocytes. Lymphocytes include T cells, B cells, and natural killer (NK) cells, which are central to identifying and combating pathogens and abnormal cells. Monocytes are another type of white blood cell that can transform into other cell types to fight infection.
To be studied, PBMCs must be separated from other blood components. This is achieved through a laboratory technique called density gradient centrifugation. In this method, a whole blood sample is layered on top of a solution with a specific density. When spun at high speed, the blood separates into layers, with the PBMCs forming a distinct band that can be collected.
The Cryopreservation of PBMCs
Cryopreservation allows for the long-term storage of PBMCs, which is important for studies that take place over many months or years. This process enables the safe transportation of samples between different laboratories. By freezing samples, researchers can collect them at various times and then analyze them together in one batch, which improves the consistency and reliability of the experimental results.
The freezing process is carefully controlled to protect the cells from damage. A step involves adding a cryoprotectant agent, such as dimethyl sulfoxide (DMSO), to the cells before freezing. This substance helps prevent the formation of ice crystals, which can rupture and destroy the cells. The cells are then cooled at a controlled, slow rate before being stored at extremely low temperatures in liquid nitrogen. This method, known as controlled-rate freezing, maintains cell viability and functionality.
Diverse Applications of Frozen PBMCs
The ability to store and thaw PBMCs has made them a resource in many areas of medical research. In immunology, they are used to study how the immune system functions and responds to various stimuli. For vaccine development, researchers analyze PBMCs from trial participants to measure the immune response generated by a vaccine candidate.
In cancer research, frozen PBMCs are used for developing and testing immunotherapies. One such area is CAR-T cell therapy, where a patient’s T cells are modified to better target and destroy cancer cells. Other applications include:
- Investigating infectious diseases by studying how immune cells react to different pathogens.
- Screening new compounds in drug discovery for their effects on the immune system.
- Identifying biomarkers that can indicate the presence of a disease.
- Predicting how a patient will respond to a specific treatment.
Handling and Assessing Frozen PBMCs
Properly handling PBMCs after they have been thawed is necessary for obtaining reliable experimental data. The thawing process must be done quickly to minimize cell stress, and the cryoprotectant agent must be carefully washed away. After thawing, it is standard practice to assess the viability of the cells to determine what percentage survived the freeze-thaw cycle.
A common method for checking cell viability is the trypan blue exclusion test. In this technique, a dye is added to a sample of the cells; dead cells absorb the dye and appear blue under a microscope, while healthy cells do not. Automated cell counters can also be used for a more standardized assessment of cell viability. Scientists also measure cell recovery, the number of viable cells obtained after thawing compared to the number frozen, to ensure sample quality before experiments.