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Cell Lines, Culture Types, & Cell Morphology
Choosing the right cell line for your experiment is an essential step in the cell culture workflow that requires careful consideration. A thoughtfully chosen cell line will support the objectives of your experiment and define other critical elements of the workflow. This includes the culture type and vessel you select as well as the morphological characteristics you will look for during culture maintenance.
Referencing the intent and objectives of your experiment before choosing a cell line can help you decide on a culture type (adherent or suspension) that can support the cell line. Once your cells are deposited in the proper vessel for your culture type, you may wish to monitor cell growth with visual inspections. Referring back to the morphology of your chosen cell line can give you an indication of the health of your cell cultures. Monitoring cellular morphology between subcultures can also help you identify early signs of contamination and irregularities in growth patterns.
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Cell lines
Choosing the proper cell line for your cell culture work relies on many experimental factors including the research intent and objective, duration of your experiment, and your requirements for consistency. Assess the goals of your experiment to choose the type of cell line that best aligns with what you want to accomplish.
In the Introduction to Cell Culture, we introduced that cell lines are derived from primary cultures following the first subculture of the isolated cells. To learn more about where cell lines come from and how they are established, please refer to that page.
If you already know your cell line, visit our Cell Culture Select Tool to search your cell line of interest and find product recommendations to help enable a successful cell culture.
Types of cell lines
Once you have determined the intent and objective, duration, and consistency requirements for your experiment, you must narrow down to a specific cell line. Consider the distinct types of cell lines with examples below.
- Species-specific: Non-human and non-primate cell lines usually have fewer biosafety restrictions, but ultimately your experiments will dictate whether to use species-specific cultures or not.
- Cell strains or tissues with functional characteristics: What is the purpose of your experiments? For example, liver- and kidney-derived cell lines may be more suitable for toxicity testing.
- Normal or transformed: Transformed cell lines usually have an increased growth rate, higher plating efficiency, are continuous, and require less serum in media, but they have undergone a permanent change in their phenotype through a genetic transformation.
- Unique growth conditions and characteristics: What are your requirements with respect to growth rate, saturation density, cloning efficiency, and the ability to grow in suspension? For example, to express a recombinant protein in high yields, you might want to choose a cell line with a fast growth rate and an ability to grow in suspension.
In addition to the criteria above, you can also choose your cell type based on:
- Is the cell line well-characterized, or do you have to perform the validation yourself?
- If you are using an abnormal cell line, do you have an equivalent normal cell line that you can use as a control?
- Is the cell line stable? If not, how easy is it to clone it and generate sufficient frozen stocks for your experiments?
The cell line you select for your experiment may have one or multiple characteristics from the list above. You may also establish your own culture from primary cells or buy established cell cultures from commercial or non-profit suppliers (i.e., cell banks). Reputable suppliers provide high quality cell lines that are carefully tested for their integrity and to ensure the culture is not contaminated. We advise against borrowing cultures from other laboratories because they carry a high risk of cell culture contamination. Regardless of their source, make sure that all new cell lines are tested for mycoplasma before you begin to use them.
Thermo Fisher Scientific offers a variety of cell lines, reagents, media, sera, and growth factors for your cell culture experiments.
Mammalian cell lines
In a cell culture lab, mammalian cell lines are the most frequently used because of their direct application to studies involving the development of human therapeutics and disease research.
These cell types require media, reagents, and culture conditions that encourage optimal growth while mimicking the right mammalian host environment.
Types of cell culture
When you have selected a cell line designed to fulfill the objectives of your experiment, you then must consider the right way to culture those cells. Most cells must be cultured while attached to a solid or semi-solid substrate. These are called monolayer or adherent cell cultures. Meanwhile, others can grow floating in the culture medium. These are called suspension cultures. The environment that the cell requires to proliferate will determine if you use an adherent or suspension culture. You may also want to consider any imaging or analysis requirements you have for the growth stage of the culture.
Table 1. Adherent vs suspension cell culture
| Adherent cell culture | Suspension cell culture |
|---|---|
| Appropriate for most cell types, including primary cultures | Appropriate for cells adapted to suspension culture and a few other cell lines that are non-adhesive (e.g., hematopoietic) |
| Requires periodic passaging, but allows easy visual inspection under inverted microscope | Easier to passage, but requires daily cell counts and viability determination to follow growth patterns; culture can be diluted to stimulate growth |
| Cells are dissociated enzymatically (e.g., Gibco TrypLE Express, trypsin) or mechanically | Does not require enzymatic or mechanical dissociation |
| Growth is limited by surface area, which may limit product yields | Growth is limited by concentration of cells in the medium, which allows easy scale-up |
| Requires tissue-culture treated vessel | Can be maintained in culture vessels that are not tissue-culture treated, but requires agitation (i.e., shaking or stirring) for adequate gas exchange |
| Used for cytology, harvesting products continuously, and many additional research applications | Used for bulk protein production, batch harvesting, and many additional research applications |
Adherent cell culture
An adherent cell culture is a culture defined by the cell’s ability to reproduce when attached to growth-promoting substrate. This characterization is called “anchorage dependence.” An anchorage dependent cell will require a plastic dish or flask that is specifically treated to allow cell adhesion and spreading, known as a tissue culture (TC) or cell culture-treated surface.
What is anchorage dependence?
Anchorage dependence is a cell’s necessity to bind to substrate to reproduce. Anchorage dependent cells require an adherent cell culture surface. Conversely, anchorage independent cells are not required to, nor can easily bind with a surface. These cells are cultured using cell culture Erlenmeyer flasks and reproduce suspended in media. Your cell’s anchorage dependence or independence will determine the type of culture plastics you will use. Choosing the proper culture vessel will ensure that your cells have a great chance for consistent cell growth.
Suspension cell culture
A suspension cell culture is one in which the cell can reproduce without tissue culture-treated vessels. They require sufficient growth medium to encourage consistent and uniform growth and occasional agitation to produce ample gas dispersion. Suspension cell cultures are passaged in shaker flasks or spinner flasks.
How to subculture suspension cells
Learn more about how to choose culture plastics
Cell morphology
Choosing the type of cell culture that is right for your cell line, whether adherent or suspension, can give your cells an excellent chance for growth. To monitor your cell’s growth, you must know the morphology and typical characteristics of your cell line.
Regularly examining the morphology of the cells in culture (i.e., their shape, structure, and appearance) is essential for confirming that your cells are healthy. Inspecting a sample of the cells by eye and a microscope each time they are handled will allow you to detect any signs of contamination early on. This will also allow you to contain contamination before it infects other cultures around the laboratory.
In some instances, your cell’s morphology will serve as the subject or variable of your experiment. Quantitative analysis of cellular morphology is also known as morphometry or morphometrics. Morphometric analysis occurs during the imaging and analysis stage of the cell culture workflow.
What is cell morphology?
Cell morphology is defined as the shape, size, appearance, structure, and function of the cell. The cell’s morphology is how we determine cell health and viability within a cell culture.
Morphological signs of deterioration of cells include granularity around the nucleus and cytoplasmic vacuolation. Signs of deterioration may be caused by a variety of reasons, including contamination of the culture, senescence of the cell line, the presence of toxic substances in the medium, or they may simply imply that the culture needs a medium change. Allowing the deterioration to progress too far can make it irreversible.
Cell shapes and sizes
Cell shapes and sizes—vital features in cell’s morphology—are uniform within the same cell line. You can identify a cell type by shape and size during imaging and analysis of cell cultures, though genetic and environmental factors may cause variation in some instances. Some cell types or strains may also be intentionally selected for diseases or mutations, such as in cancer cell culture. Variations to a cell line are available as cell strains and vary from the original cell line, therefore displaying different morphology than would be expected from the original cell line. Learn more about cell strains in the Introduction to Cell Culture.
Mammalian cells can be categorized in a few different common shapes based on the type of cell. For example, fibroblast cells, located in connective tissues, are usually flat, elongated, and spindle or star shaped [1]. Epithelial cells, from various tissues and organs in the body, can be classified as squamous with long flat shapes, cuboidal with square shapes, or columnar with a rectangular shape [2]. Lymphoblast cells appear round and spherical but can vary widely in shape.
Mammalian cell morphology
Most mammalian cells in culture can be divided into three basic categories based on their morphology.
| Fibroblastic (or fibroblast-like) cells are bipolar or multipolar, have elongated shapes, and grow attached to a substrate. |
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| Epithelial-like cells are polygonal in shape with more regular dimensions and grow attached to a substrate in discrete patches. |
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| Lymphoblast-like cells are spherical in shape and usually grow in suspension without attaching to a surface. |
 |
In addition to the basic categories listed above, if you chose a cell line with functional characteristics specific to your area of research, they may display morphological characteristics unique to their specialized role in their host. See examples below:
- Primary Human Aortic Smooth Muscle Cells (HASMC) and Primary Human Pulmonary Artery Smooth Muscle Cells (HPASMC) are elongated and sometimes triangle shaped. Most cells will have a bipolar morphology when cells reach high density in the culture, though some irregularly sized and shaped cells may be observed. Smooth muscle cells are anchorage dependent and grow best adherently.
- Neuronal cells exist in different shapes and sizes in the nervous system, but they can be divided into two basic morphological categories. Type I is defined by long axons that are used to move signals over long distances, while type II does not have axons. A typical neuron projects cellular extensions with many branches from the cell body, which is referred to as a dendritic tree. Neuronal cells can be maintained as adherent or suspension cell cultures depending on the purpose and desired outcomes of your experiment.
- Melanocytes are stellar with a multipolar or dendritic appearance and occasionally seen with small numbers of keratinocytes in the tertiary culture. In older cultures, more cells may become bipolar in appearance. Melanocytes are best maintained as adherent cell cultures.
Examples of cell lines, morphologies, and culture types
| Cell line | HEKn cells: Normal human epidermal keratinocytes isolated from neonatal foreskin and cryopreserved at the end of the primary culture stage. |
| Common applications |
|
| Recommended culture type | Adherent |
| Cell morphology | Cells are rounded, not flat, and appear in a cobblestone pattern. Cells display a high mitotic index. At near 80% confluence, the cells will be associated with each other in colonies. |
| Cell line | HUVEC: Human umbilical vein endothelial cells that have been cryopreserved at the end of the primary culture stage. |
| Common applications |
|
| Recommended culture type | Adherent |
| Cell morphology | HUVEC should have epithelioid morphology. Some irregularities in size and shape may occur but they have a cobblestone appearance with large dark nuclei and are associated with each other in colonies. During growth, cells are small and evenly sized. |
| Cell line | HDFa: Adult skin cells cryopreserved at the end of the primary culture |
| Common applications |
|
| Recommended culture type | Adherent |
| Cell morphology | Cells are refractile, spindle-shaped, and should have a bipolar morphology. They appear distributed through the culture as individual cells. |
References
- Ravikanth M, Soujanya P, Manjunath K, Saraswathi TR, Ramachandran CR. Heterogenecity of fibroblasts. J Oral Maxillofac Pathol. 2011 May;15(2):247-50. doi: 10.4103/0973-029X.84516. PMID: 22529592; PMCID: PMC3329689.
- Kurn H, Daly DT. Histology, Epithelial Cell. [Updated 2023 Feb 17]. In: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2024 Jan.
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