21/02/2023

CHO Cells

CHO Cells: From Hamsters to Heroes

Get Ready to Explore the Versatile CHO cell line in this Exciting Article!

Derived from the ovary of a Chinese hamster, the CHO cell line is a powerhouse in medical and biological research with its wide range of applications. This mammalian cell line offers endless possibilities, from recombinant protein production to gene expression, toxicity screening, nutrition, and genetic studies.

Our article delves into the fascinating world of CHO cells, exploring how these cells have revolutionized biopharmaceutical research and paved the way for life-saving therapies. Get ready to unlock the secrets of the mighty CHO cells and discover how they drive groundbreaking advances in medicine and beyond! You'll learn everything you need to know before getting started, including:

What is the CHO cell line?

CHO Cells: The Biopharmaceutical Industry's Go-To for Recombinant Protein Production
Discover the Benefits of The Mighty CHO Cells
Comparison of CHO Vs CHO-K1 cell line
Ten Tips for Culturing CHO Cells
Protocols, Videos and Recent Publications on CHO Cells

What is the CHO cell line?

Since their establishment in 1957 by Theodore T. Puck, Chinese hamster ovary (CHO) cells have become a staple in biological and medical research due to their rapid growth and high protein production. These epithelial cells, derived from the Chinese hamster's ovary, are widely used in biomanufacturing, genetics, toxicity screening, nutrition, and gene expression studies.

CHO cells can produce proteins with post-translational modifications (PTMs) similar to those found in humans. They are also deficient in proline synthesis and do not express the epidermal growth factor receptor (EGFR), making them ideal for investigating various EGFR mutations.

In biomanufacturing, CHO cells are extensively used for producing monoclonal antibodies, recombinant proteins, and vaccines. More than 60 therapeutic proteins made with CHO cells have been approved for production, and their use continues to expand. Our article looks at the remarkable properties and diverse applications of CHO cells, highlighting their crucial role in driving advances in biomedicine and beyond. Get ready to explore the fascinating world of CHO cells and discover their unparalleled potential in biomedical research!

1. CHO Cells: The Biopharmaceutical Industry's Go-To for Recombinant Protein Production

In the biotechnology industry, Chinese hamster ovary (CHO) cells are frequently used to create biopharmaceuticals like monoclonal antibodies, recombinant proteins, and vaccines.

Although you might not be aware of it, Chinese hamster ovary (CHO) cells may be to blame if you have ever undergone monoclonal antibody therapy. These adaptable cells are frequently used by the biopharmaceutical industry to produce recombinant proteins that are used in biomedical research, diagnostics, and a variety of therapeutics. Protein-based therapeutics called monoclonal antibodies (mAbs) are used to treat a variety of illnesses, such as cancer, autoimmune conditions, and infectious diseases. Because they carry out post-translational modifications resembling those in human cells, CHO cells are frequently used to make mAbs. These modifications are necessary for these therapeutics to function properly.

Proteins created through genetic engineering are known as recombinant proteins. In addition to being research reagents, they can also be used as therapeutics and diagnostics. Because they can undergo post-translational modifications and have complex glycosylations resembling those found in human cells, CHO cells are especially well suited for making recombinant proteins because of their quick growth, high protein expression, and ability to express large amounts of protein. With yields ranging from 3 to 10 grams per liter of culture, the CHO cell line is a game-changer in biopharmaceuticals thanks to its unmatched capacity to mass-produce therapeutic proteins. CHO cells are now a vital component of contemporary biomedicine thanks to genetic optimization, which increases their capacity to generate large amounts of recombinant proteins.

Vaccines are biopharmaceuticals used to prevent and treat infections brought on by viruses and bacteria. Vaccines against COVID-19 are among those made with CHO cells. Scientists have created a number of techniques, including genetic engineering, media optimization, and process development, to enhance the performance of CHO cells in the production of biopharmaceuticals. These techniques have resulted in the creation of high-yield, low-cost culture systems for the production of biopharmaceuticals using CHO cells.

The wide range of applications for CHO cells includes:

Biotherapeutics: CHO cells are used to produce various biotherapeutics, including recombinant proteins and monoclonal antibodies used in treating diseases such as cancer, autoimmune disorders, and infectious diseases.
Recombinant monoclonal antibodies: CHO cells are widely used in the production of monoclonal antibodies, which have revolutionized the field of biomedicine by providing targeted therapies for various diseases.
Active pharmaceutical ingredients (APIs): CHO cells are used to produce APIs used in the pharmaceutical industry for various therapeutic applications.

But that's not all! CHO cells have other fascinating applications in biomedical research, including:

Toxicity screening: CHO cells are used to assess the toxicity of drugs, including anti-cancer and anti-viral therapeutic agents. For example, a study explored the anti-breast cancer-specific activity of the Antarctic microalgae-derived fatty acids by using CHO as a control cell line.
Gene expression: CHO cells are used to stably and transiently express genes for gene function studies or targeted protein production. Gene editing tools are used to develop gene knock-in and knockout models in CHO cell lines.

2. Discover the Benefits of The Mighty CHO Cells

Here are some key advantages of the CHO cell line that make it an attractive research tool.

Ease of Culture: The culturing procedures and conditions of the CHO cell line are not fussy. These cells are tough and able to tolerate varying temperature and pH changes. Thus, they are ideal for large-scale culturing.
Post-Translational Modifications: These cells are similar to human cells and able to produce similar post-translational modifications. Thus, CHO cells can be used to produce biocompatible biological products with excellent pharmaceutical activity.
High productivity: CHO cells are widely used for producing high yields of recombinant proteins. Genetic optimization of the CHO cell line has resulted in approximately 3-10 grams of protein per liter of the culture.
Gene expression: CHO cells are easy to transfect; therefore, they are frequently used for transient and stable expression studies. In addition, many genetic tools are used to develop gene knock-in and knockout models using the CHO cell line.
Governmental approvals: CHO cells have been used in nearly 50 biotherapeutics approved in the USA and EU.
Low Virus Susceptibility: Due to the hamster origin, the risk of propagation of human viruses is decreased, reducing production loss and increasing biosafety.

Morphology, Cell Size and the Genome of CHO cells

Key Features of CHO Cells	Details
Morphology	Epithelial cell-like appearance, elongated and fibroblast-like shape, adherent, and grow in monolayers
Cell Size	Average 12-14 μm diameter
Genome and Ploidy	Aneuploid with 21 chromosomes compared to the euploid Chinese hamster. Karyotype shows multiple structural rearrangements and partial loss of chromosome 2 and X material is detected.

Microscopic images of CHO cells: at high confluency (left) and at around 50% confluency (right).

3. Comparison of CHO Vs CHO-K1 cell line

Since the original CHO cell line was reported in 1956, many variations of the cell line have been created for various purposes. CHO-K1 was generated from a single clone of CHO cells in 1957, and CHO-DXB11 (also known as CHO-DUKX) was subsequently made through mutagenesis with ethyl methanesulfonate. However, their utility was limited due to their ability to revert to DHFR activity when mutagenized. Later, CHO cells were mutagenized with gamma radiation to produce CHO-DG44, in which both DHFR alleles were entirely eliminated. These DHFR-deficient strains require glycine, hypoxanthine, and thymidine for growth and are widely used for industrial protein production. Other selection systems have since become popular, and host cells such as CHO-K1, CHO-S, and CHO-Pro minus have been shown to produce high levels of proteins. Due to genetic instability, these cell lines are often cultivated in animal component-free or chemically defined media in suspension culture bioreactors. The complexities of CHO cell genetics and clonal derivation were also discussed.

4. Ten Tips for Culturing CHO Cells

The CHO cell line is a low-maintenance cell line that is easy to culture.
CHO cells have a fast population doubling time of 14-17 hours.
CHO cells are adherent and grow as monolayers.
Subculture CHO cells at 80-90% confluency using Accutase.
Seed CHO cells at 1 x 10⁴ cells/cm2 cell density to yield a confluent monolayer in around 4 days.
For optimal culturing, use a 50:50 DMEM and Ham's F12 mixture supplemented with 5% FBS and L-glutamine.
Renew the growth medium 2-3 times a week.
Cultivate CHO cells in a humidified incubator supplemented with 5% CO2 gas at 37°C.
Store CHO cells in liquid nitrogen's vapor or liquid phase (-196°C).
Follow the Biosafety level 1 guideline for handling and culturing the CHO cell line.

5. Protocols, Videos and Recent Publications on CHO Cells

Here are some excellent resources to explore for learning about CHO cell line culturing and maintenance.

An extensive cell culture protocol on CHO cells: This link can help you learn all about CHO cell subculturing and transfection.
CHO cells: This site will provide basic cell culture information about the CHO cell line, including splitting, storage, freezing, and thawing of cells, etc.
Thawing CHO cells: This video shows an exemplary thawing protocol for frozen CHO cells.

5.1. Transfection protocols of CHO cell line

CHO cells are highly amenable to both transient and stable transfection of genes. Here are some resources providing helpful information about CHO cell line transfection protocols.

CHO cell transfection: This published article provides a transient transfection protocol for the CHO cell line using linear polyethyleneimine (PEI).
Transfection methods for CHO cells: This article explains different strategies for CHO cell line efficient transfection using different transfection reagents.
Transient transfection of CHO cells: This video uses illustrations to explain basic concepts regarding transient expression studies in CHO cells.

5.2. Interesting Research Publications on CHO Cells

Study	Year	Purpose	Methodology	Key Findings
Rapid, high-yield production of full-length SARS-CoV-2 spike ectodomain by transient gene expression in CHO cells	2021	Expression of SARS-CoV-2 spike ectodomain in CHO cells using three transient transfection methods for high productivity	CHO cells were transfected with plasmids encoding the full-length SARS-CoV-2 spike ectodomain using three transient transfection methods. Protein expression was assessed by ELISA and Western blot.	All three transient transfection methods showed high levels of protein expression, with the highest yield obtained by the polyethylenimine method.
Engineering a stable CHO cell line for the expression of a MERS-coronavirus vaccine antigen	2018	Production of MERS-coronavirus antigen in CHO cells for use as a future candidate vaccine	CHO cells were transfected with a plasmid encoding the MERS-coronavirus antigen and selected for stable expression using geneticin. Protein expression was assessed by ELISA and Western blot.	The stable CHO cell line showed high levels of protein expression and stability over multiple passages.
Cytotoxic activity of fatty acids from Antarctic macroalgae on the growth of human breast cancer cells	2018	Use of CHO cells as a control to assess the toxicity of anti-cancer agents against normal cells	CHO cells were cultured and treated with fatty acids from Antarctic macroalgae, and cell viability was assessed using MTT assay.	Fatty acids from Antarctic macroalgae showed no cytotoxic effects on CHO cells, suggesting potential use as an anti-cancer agent with selectivity for cancer cells.
Knockout of caspase-7 gene improves the expression of recombinant protein in CHO cell line through the cell cycle arrest in G2/M phase	2022	Genetic manipulation of CHO cells to improve the expression of recombinant proteins	Caspase-7 gene was knocked out in CHO cells using CRISPR/Cas9 technology, and protein expression was assessed by Western blot and fluorescence microscopy.	Knockout of caspase-7 gene in CHO cells resulted in improved protein expression, likely due to the G2/M phase cell cycle arrest caused by the loss of caspase-7.
Development of a CHO cell line for stable production of recombinant antibodies against human MMP9	2015	Production of monoclonal antibodies against human MMP9 protein in CHO cells	CHO cells were transfected with plasmids encoding the antibody against human MMP9 and selected for stable expression using geneticin. Protein expression was assessed by ELISA and Western blot.	The stable CHO cell line showed high levels of antibody expression and stability over multiple passages, suggesting potential use in therapeutic applications targeting human MMP9.

We hope the information in this article will help you before starting with the CHO cell line. If you consider using this cell line in your research study, consider ordering from us!

References

Reinhart, D., et al., Bioprocessing of recombinant CHO‐K1, CHO‐DG44, and CHO‐S: CHO Expression hosts favor either mAb production or biomass synthesis. Biotechnology journal, 2019. 14(3): p. 1700686.
Pan, X., et al., Metabolic characterization of a CHO cell size increase phase in fed-batch cultures. Applied microbiology and biotechnology, 2017. 101: p. 8101-8113.
Turilova, V.I., T.S. Goryachaya, and T.K. Yakovleva, Chinese hamster ovary cell line DXB-11: chromosomal instability and karyotype heterogeneity. Molecular Cytogenetics, 2021. 14(1): p. 1-12.
Hunter, M., et al., optimization of protein expression in mammalian cells. Current protocols in protein science, 2019. 95(1): p. e77.
Nyon, M.P., et al., Engineering a stable CHO cell line for the expression of a MERS-coronavirus vaccine antigen. Vaccine, 2018. 36(14): p. 1853-1862.
Pacheco, B.S., et al., Cytotoxic activity of fatty acids from Antarctic macroalgae on the growth of human breast cancer cells. Frontiers in Bioengineering and Biotechnology, 2018. 6: p. 185.
Ryu, J., et al., development of a CHO cell line for stable production of recombinant antibodies against human MMP9. BMC biotechnology, 2022. 22(1): p. 8.