CHO-K1 Cells: A Staple in Biotechnological Research Applications

CHO-K1 cells are derivative of the Chinese hamster ovary (CHO) cell line. They are extensively used in industrial biotechnology to produce biopharmaceutics and other recombinant proteins. Besides, the CHO-K1 cell line is also employed in toxicology research. Researchers genetically manipulate these cells to improve glycosylation, reduce apoptosis and upsurge overall productivity.

This article will help you go through almost all the valuable knowledge about the CHO-K1 cell line that can ease your work with it. Specifically, it will cover:

Origin and general characteristics: CHO-K1 cells
CHO-K1 cell line: Culturing information
Advantages & Limitations of CHO-K1 cells
Applications of CHO-K1 cell line in research
CHO-K1 cells: Research publications
Resources for CHO-K1 cell line: Protocols, Videos, and More

1. Origin and general characteristics: CHO-K1 cells

A cell line's general characteristics and origin determine its use in research. This section will help you learn about the origin and features of the famous CHO-K1 cell line. You will know: Where did the CHO-K1 cell come from? How big are CHO-K1 cells? What is the full form of the CHO-K1 cell line? What is the CHO-K1 cell morphology?

CHO-K1 or Chinese hamster ovary cell line K1 is a subclone of parental CHO cells that originated from the biopsy of an adult female Chinese hamster ovary in 1957. The original cell line was developed by T.T. Puck and colleagues at the University of Colorado Medical School, Denver, USA [1].
CHO-K1 cell line exhibit epithelial-like morphology.
The diameter of CHO-K1 cells is approximately 0.001 millimeters. Interestingly, the cells are big initially; however, they get small over time.
The CHO-K1 genome consists of a similar number of chromosomes as human cells. They have diploid karyotypes and have only fewer chromosomal abnormalities.

CHO-K1 vs CHO-S cell line

Both cell lines are CHO derivatives. They differ in the way these cells grow and proliferate. CHO-S cells are adapted to grow in cultures, whereas CHO-K1 can be genetically manipulated to produce adherent and suspension cells.

Suspension culture in pharmaceutical production.

2. CHO-K1 cell line: Culturing information

The CHO-K1 cell line is widely used in industrial biotechnology research. They are easily maintainable cell lines. Knowing about the CHO-K1 cell culturing key points can help you go easy with it. This section will assist you in learning: Are CHO-K1 cells adherent? What is the doubling time of CHO-K1 cells? What media is used for CHO cell culture? What is CHO-K1 seeding density?

Key Points for Culturing CHO-K1 Cells

Population Doubling Time:	The CHO-K1 doubling time is approximately 22 hours.
Adherent or in Suspension:	CHO-K1 cells are adherent. However, they can be genetically modified to be CHO-K1 suspension cells.
Seeding Density:	CHO-K1 seeding density is 1 x 10⁴ cells/cm². At this density, cells can make a confluent layer in approximately 6 days. For adherent cells, cells are rinsed with 1 x PBS and incubated for 8 to 10 minutes at ambient temperature. Dissociated cells are added to fresh medium and centrifuged. Harvested cells are resuspended and poured into the new flask for growth.
Growth Medium:	Ham's F12 growth medium supplemented with 10% FBS, 1.0 mM stable Glutamine, 1.0 mM Sodium pyruvate, and 1.1 g/L NaHCO3 is used to culture CHO-K1 cells. Media should be replaced 2 to 3 times a week.
Growth Conditions:	CHO-K1 cultures are kept in a 37°C humidified incubator with a 5% CO2 supply.
Storage:	Frozen CHO-K1 cells are stored at below -150°C temperature or in the vapour phase of liquid nitrogen.
Freezing Process and Medium:	CM-1 or CM-ACF freezing media are used for CHO-K1 cell freezing. A slow freezing process that allows a gradual 1°C drop in temperature is used to freeze CHO-K1 cells.
Thawing Process:	Frozen CHO-K1 cells are kept in a 37°C water bath until a small ice clump is left. Thawed cells are added with fresh culture media and poured into the new flask containing culture media at a density of 5 x 10⁴ cells/cm². Cells take almost 24 to 48 hours to revive appropriately.
Biosafety Level:	CHO-K1 cultures are handled and maintained in biosafety level 1 laboratories.

CHO-K1 cells in suspension culture showing less than 10% adherent cells (left) and clustering (right).

3. Advantages & Limitations of CHO-K1 cells

CHO-K1 is an invaluable research tool. Its unique combination of advantages and limitations makes it distinguishable from other cell lines. This section has discussed a few pros and cons of the CHO-K1 cell line.

Advantages

The main advantages of the CHO-K1 cell line include:

Transfection amenability	CHO-K1 cells are widely used in transfection studies. They can be transfected transiently and stably through different physical and chemical procedures. Due to their high transfection amenability, CHO-K1 cells are extensively employed to produce recombinant proteins and other biopharmaceuticals.
Fast growth rate and easy culturing	CHO-K1 cell doubling time is only 22 hours, thus, they have a high growth rate and are ideal for industrial biotechnology use. Moreover, CHO-K1 suspension adaptation makes them useful for producing large quantities of biopharmaceuticals. Besides, they are easy to culture and maintain in laboratories and require no difficult culturing conditions and procedures.
Low frequency of chromosomal abnormalities	CHO-K1 is a well-characterized and established model system. The CHO-K1 genome is stable and has only fewer chromosomal abnormalities. Therefore, they are ideal hosts for the production of recombinant proteins.

Limitations

Here are some limitations of the CHO-K1 cell line:

Non-human origin	Although CHO-K1 cells have the ability to carry out human-like glycosylation patterns, they are of non-human origin. This might be a concern when studying highly human-specific cell processes and the immunogenicity of therapeutic agents.
Heterogeneity	CHO-K1 cells may exhibit slightly different genetic characteristics within the same population, resulting in genetic heterogeneity. This may affect cellular functions and cause variability in protein expression levels, potentially influencing the reproducibility of experimental results.

4. Applications of CHO-K1 cell line in research

The CHO-K1 cell line has numerous applications in industrial biotechnology and toxicology research. Here, we have discussed some specific ones.

Recombinant protein production: CHO-K1 cells are invaluable research tools for the production of recombinant proteins, including antibodies, therapeutic proteins, and enzymes. Their high growth rate and easy culturing conditions help to produce large quantities of recombinant proteins with proper folding and glycosylation. Such as, a study conducted by Kritika Gupta used CHO-K1 cells and stably transfected them to produce a recombinant monoclonal antibody against tumour necrosis factor-alpha (TNF-α) [2]. CHO K1 antibody production is quite reliable and convenient. Researchers also modify these cells for improved CHO K1 antibody production. For instance, a study genetically manipulated CHO-K1 cells to produce antibodies with high ratios of a-fucosylated Fc-attached N-glycans, which is important for their effector function [3].
Toxicology research: The CHO-K1 cell is often used for drug discovery and screening assays. They can be employed to evaluate the toxicity and efficacy of potential drugs. Moreover, researchers use CHO-K1 cells to investigate drug-target interactions and study drug metabolism. Several studies have been carried out to assess the possible therapeutic effects of plant extracts, compounds, nanoparticles, therapeutic proteins, and other agents using the CHO-K1 cell line. A similar research was conducted in 2022 in which researchers measured the cytotoxic potential of flavonoid-rich plant extracts in CHO-K1 cells [4]. Likewise, a research study conducted by Ryan Deweese and colleagues evaluated the cytotoxicity of Baptisia australis, Trifolium pratense, and Rubus idaeus extracts on chine hamster ovary CHO-K1 Cells [5].

5. CHO-K1 cells: Research publications

The following are some interesting research publications on CHO-K1 cells.

Overexpression of SIRT6 alleviates apoptosis and enhances cell viability and monoclonal antibody expression in CHO-K1 cells

This study published in Molecular Biology Reports (2023) proposed the positive effects of SIRT6 gene overexpression on CHO-K1 cell viability and antibody expression.

Enhancing the yield and activity of defucosylated antibody produced by CHO-K1 cells using Cas13d-mediated multiplex gene targeting

This publication is in the Journal of the Taiwan Institute of Chemical Engineers (2021). The research findings state the potential of CRISPR-Cas13d for genetically modifying CHO-K1 cells for improved antibody production concerning quality and quantity.

Application of maltose as energy source in protein-free CHO-K1 culture to improve the production of recombinant monoclonal antibody

This research article from Nature Scientific Reports (2018) suggested maltose as a promising energy source for culturing CHO-K1 cells in a protein-free medium and enhancing recombinant monoclonal antibody production.

Reveal cytotoxicity and antigenotoxicity of piper nigrum l. ethanolic extract and its combination with doxorubicin on CHO-K1 cells

This study in the Indonesian Journal of Cancer Chemoprevention (2018) used CHO-K1 cells to evaluate the potential cytotoxic and antigenotoxic effects of black pepper ethanolic extract alone and in combination with doxorubicin.

Cytotoxicity and genotoxicity of silver nanoparticles in Chinese Hamster ovary cell line (CHO-K1) cells

This research was published in The Nucleus in 2019. Herein researchers assessed silver nanoparticles' cytotoxic and genotoxic potential in the CHO-K1 cell line.

6. Resources for CHO-K1 cell line: Protocols, Videos, and More

CHO-K1 is a famous cell line. The available resources featuring CHO-K1 culturing and transfection protocols are mentioned here.

CHO-K1 transfection: This link will describe the CHO-K1 transfection protocol. It is a step-by-step guide for transfecting plasmid DNA into CHO-K1 cells using Lipofectamine LTX Reagent.
CHO-K1 transfection tutorial: This video will explain the transient CHO-K1 transfection procedure in detail.

Here are some resources describing cell culture protocol for CHO-K1 cells.

CHO-K1 cells: This website link contains useful information about CHO-K1 cells, including CHO-K1 media recipe, subculturing, and thawing protocol.

References

Gamper, N., J.D. Stockand, and M.S. Shapiro, The use of Chinese hamster ovary (CHO) cells in the study of ion channels. J Pharmacol Toxicol Methods, 2005. 51(3): p. 177-85.
Gupta, K., et al., A Stable CHO K1 Cell line for producing recombinant monoclonal antibody against TNF-α. Molecular Biotechnology, 2021. 63(9): p. 828-839.
Popp, O., et al. Development of a pre-glycoengineered CHO-K1 host cell line for the expression of antibodies with enhanced Fc mediated effector function. in MAbs. 2018. Taylor & Francis.
Kurchatova, M., et al., Cytotoxicity of Flavonoid-Containing Plant Extracts toward the CHO Cell Line: a Comparative Study. Cell and Tissue Biology, 2022. 16(1): p. 80-85.
Deweese, R., et al., Cytotoxic Effects of Trifolium pratense, Baptisia australis, and Rubus idaeus Extracts on CHO-K1 Cells. GSC Advanced Research and Reviews, 2021. 8(1): p. 128-139.