At BioIVT, our wide breadth of isolated cells allows us to touch many different areas of research. From oncology to autoimmune and everywhere in between, our capabilities cover a variety of disease indications and research applications. A struggle that researchers sometimes face is which cells to use in their studies and when to use them. Primary cells, dissociated tumor cells, immune cells, and cell lines all have their place in research. Some overlap but also have their own niche.
Immortalized cell lines have been a popular research tool for over 50 years, and while they remain useful for researchers, advances in technology have allowed for the development of methods to isolate and culture other cell types that may be more appropriate for your research.
What exactly are these different cell types? How are they derived and how do they relate to each other? For starters, all cell types originate either from tissue or biofluids. There are four main types of cells:
DTCs – Dissociated Tumor Cells (DTCs) are derived from fresh tumor tissue, which is dissociated into a single cell suspension. The cells are then cryopreserved without any further manipulation.
Immune cells – Immune cell subpopulations can also be isolated from tissue or biofluids. Various sub-populations can be isolated using various techniques and depend on the amount of specific subsets in the starting material.
Primary Culture Cells – If tissue is dissociated into a single cell suspension and then the cells are placed into culture instead of being cryopreserved, the resulting cell population is generally composed of fibroblasts or epithelial cells. These cells are frequently termed “primary” cells as they will grow in culture for a few passages but are not immortalized. Immune cells and primary culture cells can be transformed into immortalized cell lines.
Immortalized Cells
Immortalized cells can be propagated in culture indefinitely - a very useful quality - especially when one is looking to use the same cells over and over in multiple assays. Well-established cell lines also often have known genetic profiles and are well characterized. However, the main drawback of cell lines is that at least compared to these other cell types, they are very far from the original source material, usually having been intentionally modified so they may not be representative of in vivo conditions.
To derive DTCs, the tissue is first minced and then put through enzymatic and further mechanical digestion. The suspension is then strained, washed and frozen. Since DTCs come from the dissociation of a fresh piece of tumor tissue, they will be a heterogeneous mixture of all the cells present in the tumor and may contain tumor infiltrating lymphocytes (TILs).
DTC products need to be well characterized, and quality checked. First, it’s important to have the original tumor tissue reviewed by a pathologist to confirm the tissue type and disease state. Sterility, mycoplasma, and human pathogen free testing are important for downstream uses, particularly culturing. STR profiling is used to confirm that there has been no contamination of these samples with known cell lines. Performing a post-thaw cell count, viability assessment, and growth assessment can give researchers an idea of how the cells perform in downstream applications. To characterize DTCs, it is helpful to look at the expression of common leukocyte markers. This data will vary with each tumor and is useful for researchers interested in TILs and can indicate the presence of certain leukocyte subsets.
DTCs will vary widely from tumor to tumor, and the cells from different tumors or donors will perform differently in culture. Some will obtain a fibroblast-like morphology in culture while others will appear more epithelial-like. Many will seem to be mixed cultures while others won’t proliferate in culture at all. The most important thing to remember about DTCs is that they provide the closest known comparison to the original tissue while still offering the flexibility of working with cells. Another factor to consider when looking at DTC products is what kind of clinical data set is available from the DTC donor. More detailed clinical data results in more variables to analyze. One more aspect of DTC products that may be important to researchers is the availability of matched PBMCs or cells from matched normal adjacent tissue.
While DTCs are frozen down after dissociation, straining and washing, primary cells are instead put into culture. The type of cells that arise will depend on the starting tissue, with many of these cultures generally become fibroblastic or epithelial-like in appearance. The longer primary cells are in culture, the more phenotypic drift they will undergo, making them less representative of clinical conditions and less valuable to researchers. Therefore, primary cells used after only 1 or 2 passages are best.
Primary cells have a slower growth rate than cell lines. They are generally passaged about every two weeks instead of every few days. While primary cells are often generated from tumor tissue, cells from other tissue types can be used also and researchers may be particularly interested in primary culture cells from diseased tissues like IPF and CF lungs or RA and OA synovium.
Primary cells should generally be quality checked and characterized in the same way as DTCs, with pathology review, contamination testing, and post-thaw growth checks all being important steps. Since primary culture cells often appear to be fibroblasts or epithelial cells, performing a leukocyte marker expression assessment is not as useful for them. Instead, primary culture cells can be assessed for the expression of a fibroblast marker, such as vimentin, or for an epithelial cell marker, such as cytokeratin, to confirm their fibroblast or epithelial cell type.
Primary culture cells are more purified populations than DTCs because culturing often leads to selective expansion of fibroblasts or epithelial cells over the other cell types present. When these primary culture cells come from oncology tissues, they are referred to as cancer-associated fibroblasts (CAFs) and cancer-associated epithelial cells (CAEs), when they come from diseased or normal lung tissue they are referred to as Primary Parenchymal Fibroblasts or Primary Bronchial Epithelial Cells. Primary culture cells are quite similar to the cells in the original tissue, especially if they’ve only been cultured for a short time. Like DTCs, the availability of matched normal cells and PBMCs along with the presence of a complete clinical data set, quality check, and characterization data are all important factors to consider when looking at primary cell products.
Because of the critical role the immune system plays in disease progression, researchers studying a wide variety of diseases will require immune cells from relevant donors. Immune cells are frequently isolated from a variety of tissue types - most commonly peripheral blood, bone marrow, cord blood, and spleen/lymph node or tonsil tissue. These isolations are often performed by density gradient centrifugation, which yields a population of mononuclear cells (MNCs). A wide variety of cell types can be isolated from the mononuclear cells, including T and B cells, NK cells and monocytes. Hematopoietic stem cells can be generated from cord blood and bone marrow MNCs.
Samples for immune cell isolation can be collected in a variety of formats including whole blood, buffy coats, CPT tubes, and leukopaks.
- Whole blood is most common and regularly collected in all biomedical settings. Different anticoagulants can be chosen based on specific requirements.
- Buffy coats are generated from whole blood units that are enriched for MNCs.
- CPT tubes have a gel barrier that allows for quick and easy PBMC isolation.
- Leukopak samples are collected by leukapheresis and have a very high yield of MNCs from a single donor.
Cell Applications
Cell products are valuable for use in a variety of applications. Cell products might be used in biomarker discovery studies, especially those from a wide variety of disease types that may not be accessible in other formats. Cells that best represent in vivo conditions will be the most relevant for these kinds of studies, making DTCs and primary cells the most appropriate cell product for biomarker discovery research. Cell products are also useful in drug discovery research. It’s critical to have the appropriate disease type for candidate compound screening assays, making a diverse inventory of diseased cell products invaluable to researchers.
Cell therapy is an up and coming area of research, and because DTCs, primary cells, and immune cells come directly from human donors with no modification, they are valuable to cell therapy researchers who might use them as targets for cell killing assays or to validate gene editing methods.
Cells from diseased tissue are often necessary for diagnostic development work, and thus access to a wide variety of disease types including oncology, respiratory disease, and joint disease is valuable to those developing new diagnostic methods.
Immunotoxicity is an important factor in medical device validation. Access to a large and diverse inventory of immune cells from a wide breadth of disease indications, as well as from different species, is a useful tool for those assessing the immunotoxicity of medical devices. Cell lines, primary cells, and some DTCs can be incorporated into 2D and 3D culture models. These viable human cells will provide a better representation of the clinical environment, helping to generate more relevant models for drug screening and chemosensitivity testing.
Personalized medicine researchers can use BioIVT’s patient-derived cells to determine how specific disease cohorts will react to different drug therapies, as our cell products (especially our DTCs) provide close representations of the clinical scenario. While no two patients’ cell populations are identical, there is a greater likelihood of therapy being effective if it has been tested on cells from donors with similar diagnoses.
Our large clinical sourcing network gives us the ability to gather the tissues required to get you the cells you need. Through our recent acquisitions of three donor centers in Pennsylvania, we now have access to reliable, recallable leukopak donors, from which we can provide a high yield of immune cells from the same donor. For more information regarding BioIVT’s wide-breadth of cell types, please click here.
