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Utilizing Primary Cell Lines and 3D Cell Models to Better Represent a Living System

From transformed cell lines to primary cell lines and 3D modeling, there is a cell model for every application

Marnie Willman

Marnie Willman is the clinical writer at Today's Clinical Lab. She can be reached at

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Cell models and tissue culture form the epicenter of critical research worldwide, used for testing new therapies, modeling disease, and exploring the unknown. Science writer Marnie Willman spoke to Chongbei Zhao, MD, PhD, the head of Cells, Tissues, and Organoids Center at Stowers Institute for Medical Research. She explains some of the considerations surrounding different cell models and use of tissue culture in medical research. Note: These responses have been edited for clarity and style.

Portrait of Chongbei Zhao
Chongbei Zhao, MD, PhD
Credit: Chongbei Zhao

Q: What are the benefits and limitations of using cell lines for research, compared to animal models or other approaches?

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A: Cell lines are well established under specific lab conditions, making them excellent models for various studies. Compared to housing and maintaining animals, using cell models is much less expensive and technically easier to handle. Cell cultures also do not come with the ethical considerations that are a mandatory component of animal research. Cell models have been used for over 100 years, and there is a wide variety of cell lines openly accessible for a wide range of research. 

New options such as 3D cellular models and organ-on-a-chip systems are beginning to break down the barrier of lacking physiological complexity, which was originally one of the limitations of cellular models. Another predominant limitation of cell models is that genetic variability can occur as the cell lines are maintained over time. These accumulated changes can potentially alter their properties, affecting reproducibility of experimental results. 

Q: Your group works with several different primary cell lines. Can you explain what primary cell lines are and how they’re different? 

A: Primary cell lines are isolated and derived directly from a living organism, unlike other types of cell lines like immortalized cells. Immortalized cell lines have been manipulated to continue replicating and dividing indefinitely. Because primary cell lines aren’t manipulated this way, they more accurately represent the physiological properties of cells from the living organism being modeled. When modeling metabolism, aging, signaling, and other physiological processes, primary cell lines are a more representative model than immortalized cell lines. 

A common misunderstanding is that cell line-based research findings are directly translatable to animal models or human physiology.

The difficulty in working with primary cell lines is that they may require additional nutrients and specific experimental conditions (such as adjusted temperature, pH, environmental oxygen, etc.) to replicate. They also do not replicate indefinitely, and after a certain number of cell divisions, they undergo senescence. Primary cell lines may be less accessible than well established, immortalized cell lines, and can require specialized techniques or protocols to handle in a laboratory setting. 

Q: How does your research group use long-term primary cell lines?

A: We use both 2D and 3D models as 3D culture conditions provide a better culture system for understanding cell biology, biochemistry, cancer biology, drug screening, and modeling neurodegenerative diseases. We have also used gene editing to transform primary cell lines into immortalized models for disease modeling, which is a very useful application. We derived primary mouse fibroblast cells for cell biology studies to support the growth of embryonic stem cells or induced pluripotent stem cells, which are used in several stem cell projects we’re involved in. 

Q: What are some of the challenges you’ve experienced working with primary cells?

A: While primary cells provide a better model for our work, they do present some complications. We work with many non-traditional model organisms including apple snails, planarians, nematostella, and cavefish. These are susceptible to contamination by bacteria, mold, yeast, and mycoplasma. These primary cells also have higher requirements for culture conditions and have a more limited lifespan than immortalized cell lines. Sometimes, fresh tissues or the quantity of tissues from the desired animal model are not available, which can delay or stall the research. We also must always consider that primary cells may exhibit heterogeneity within the cell population and take these differences into account when interpreting the data. 

Q: What do you think is misunderstood about cell line-based research? 

A: Whether working with primary cells or an immortalized line, a common misunderstanding is that cell line-based research findings are directly translatable to animal models or human physiology. It’s important to keep in mind that cell lines are simplified models of complex biological systems. They are cultured in a dish without intricate interactions between tissues, organs, and physiological cues. While 3D cell models aim to improve this, cell models do not perfectly represent the living organism. While maintaining cell lines, genetic mutations over time can occur regardless of what type of cells are being used. Immortalizing cells to allow indefinite growth can also affect cellular properties and introduce differences from the original tissues. When cell lines are derived from disease tissues, such as tumors, they may not fully recapitulate the complexity and heterogeneity of human diseases. Cell line models are an essential stepping stone during initial discovery and characterization, when too little is known to justify animal research.  

Chongbei Zhao graduated from Zhengzhou University in China with her MD. She earned her PhD in cell biology from the University of Missouri in Columbia. Chongbei is currently the head of the Genome Engineering Facility, Media Preparation Facility, and the Cells, Tissues, and Organoids Center at Stowers Institute for Medical Research in Kansas City, Missouri. Her current research focuses on cell biology, primary cell line development, media optimization, organoids development, and gene editing (CRISPR/Cas) technology development with various species.