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Peripheral Blood Mononuclear Cells — What You Should Know?

Peripheral Blood Mononuclear Cells (PBMCs) are any blood cell with a single nucleus. These blood cells are a critical component in the human immune system to fight pathogens. PBMCs consist of lymphocytes (T, B, and NK cells), monocytes, and dendritic cells. Peripheral blood mononuclear cells (PBMC) are hematopoietic stem cells (HSC) that develop in the bone marrow and are an important line of defense against disease and infection in the peripheral blood. PBMCs are widely used in research fields including immunology, infectious diseases, hematological malignancies, etc.

Bio-Sample Sources

The most common starting material for PBMC isolation is peripheral blood. However, PBMC can also be isolated from other blood products, including whole blood, buffy coat, leukoreduction system chamber, leukopak, and others.

Bio-sample Description Concentration of PBMCs
Whole blood (WB) WB is human blood from standard blood donations and consists of red blood cells, white blood cells, and platelets. The number of PBMCs per milliliter of blood can vary depending on the patient’s age. 0.5-3x106 PBMCs/mL
Buffy coat A "Buffy Coat", also known as leukocyte concentrate, is a byproduct in manufacturing red blood cell and platelet concentrate from a whole blood donation. There is a much higher concentration of PBMCs in these samples. ~1x107 PBMCs/mL
Leukoreduction system chamber Leukoreduction system chambers can be used to isolate PBMCs at a high concentration. 0.6-1x108 PBMCs/mL
Leukopak A Leukopak is a highly-enriched leukapheresis-derived product. ~5x107 PBMCs/mL

Methods for Isolating PBMCs

PBMC isolation protocols involve the isolation of lymphocytes directly from biological samples such as whole blood. PBMCs occupy only a small portion of a biological sample and are extremely difficult to study when crowded with other substances. Therefore, researchers must first isolate PBMC from whole blood before they can perform toxicology experiments.

  • Density gradient medium
    PBMCs, which include lymphocytes and monocytes, have a specific gravity of about 1.070, whereas erythrocytes and polymorphonuclear leukocytes have a specific gravity of more than 1.080. This allows for density gradient centrifugation using a solution between the two specific gravities (called the stratification solution) to obtain single nucleated cells.
  • Frit barrier
    Tubes containing a frit barrier provide better separation of the blood layers during the centrifugation step. The use of a frit barrier decreases the overall processing time and improves standardization.
  • Cell preparation tubes (CPT)
    To provide easier PBMC isolation, CPT contains an anticoagulant plus a gel product that aids blood layer separation. CPT tubes can be centrifuged on-site and then shipped to the laboratory for the final isolation steps.
  • Immunomagnetic cell separation
    Immunomagnetic cell separation (column-based or column-free) is a method for providing high-yield, high-purity PBMCs. This technique can also be used to enrich a sample for specific immune cell subsets.

Counting PBMCs

The final step in the isolation process is to assess PBMC quality. The viability and quantity of PBMCs can be estimated via manual cell counting. However, manual cell counting can be time-consuming and result in variation between operators. Automated counters provide an alternative standardized approach that is quicker to perform and often the preferred method for clinical research studies.

Freezing and Thawing PBMCs

  • Freezing process
    Cryopreservation is a commonly used method for the long-term storage of cells, including human peripheral blood mononuclear cells (PBMCs) isolated from whole blood or leukopaks. To cryopreserve PBMCs, the cells are resuspended in a cryopreservation medium, cooled to extremely low temperatures, then stored at liquid nitrogen temperatures (below -135°C) until needed.
  • Thawing process
    When the cells are needed, the frozen PBMCs can be thawed. Appropriate techniques and handling ensure optimal cell viability, recovery, and function in downstream applications.

PBMCs in Clinical Research

Over the past 50 years, PBMC has become increasingly important in clinical research. One reason for its growing popularity is the need for PBMC to develop CAR-T cells for cell and gene therapy. Other uses include, but are not limited to patient stratification, biomarker identification, immuno-oncology, toxicology, and rare disease research.

Creative Bioarray Relevant Recommendations

Product Types Description
CD4+ T cells Coordinate adaptive immunity through activation and regulation of other immune cells
CD8+ T cells Destroy cancer cells or cells that are infected or damaged
B cells Secrete antibodies as part of the humoral immune response
NK cells Trigger lysis or apoptosis of infected cells
Monocytes Take up foreign objects via phagocytosis, perform antigen presentation, and produce cytokines
Dendritic cells Process and present antigen material to T cells
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