Common Problems in Cell Culture

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Cell culture is the process of isolating cells from an organism and growing them in a scientifically controlled environment. Contamination is a common problem in cell culture, and contaminants include mycoplasma, yeast, bacteria, and chemicals.

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Origins of cell culture

The practice of cell culture began as tissue culture in the year 1907 when Ross Harrison, an American biologist and pioneer in experimental biology, developed the “hanging drop technique”.

Harrison removed the nerve fibers of a frog and placed them in a salt medium containing serum. He observed that the cells were able to easily grow in the new environment. This was the beginning of animal cell culture. Since Harrison’s discovery, there have been several advances in cell culture techniques.

Types of cell culture

The different cell culture techniques are organ culture, adherent cell culture, suspension culture, primary culture, and continuous cell lines.

Organ culture

In organ culture, the entire organ is explanted, preserving cell-cell interactions. Organ culture is difficult to use for quantitative determination, as the variation between different experimental batches is higher.

Adherent cell culture

Adherent cell culture involves the growth of cells as adherent monolayers. The cells are dependent on the substratum (basal layer) for proliferation, hence adherent monolayers stop dividing once two cells come into contact with each other.

Suspension culture

In a suspension culture, the bond between the cells or between the cells and the substratum are broken down using proteolytic enzymes for further harvesting.

Primary culture

Primary cultures involve the isolation of cells directly from the animal.

These cells ultimately die and a new tissue needs to be obtained from the animal once again for preparing a new culture. This is because cells that divide in culture will eventually cease to divide upon repeated subculturing, reflecting the process of senescence that usually occurs to all cells in situ.

Continuous cell lines

To counter the issue of senescence, continuous cell lines were developed. These cell lines are usually obtained by culturing tumor cells and do not require any substratum for their growth.

Common problems in cell culture

There are many practices for handling cell lines, cell culture media and reagents to avoid cross-contamination. Some of the common issues faced are stated below:

Contamination by bacteria, fungi, and yeast

This type of contamination can happen through laboratory equipment, reagents, and even through the operator’s hair, hands, breath, or clothing. Particulate fallout or aerosol fallout during any stage of culturing will lead to biological contamination.

In addition, laboratory facilities such as laminar air flow cabinets, carbon dioxide humidified incubators, and water baths are sources for biological contamination apart from contamination caused due to dust, aerosols, or presence of crevices. Flask with loose caps and improperly sealed culture plates pave way for contaminants to crawl, swim, and grow in the cultures.

Mycoplasma contamination

Known as the “crabgrass” of cell cultures, mycoplasmas have the ability to alter the cell function of the host culture bringing about cytopathic effects.

Chemical contamination

Wrong constitutions of reagents or substandard quality of water, sera, or containers used to make the reagents are some reasons for chemical contamination of a cell culture.

Endotoxins, the byproduct of gram-negative bacteria, are sometimes found in sera and other additives that affect the growth of a culture.

Preventing culture contamination

The following steps can be taken to reduce the risk of culture contamination:

  • Use of good aseptic techniques;
  • Reducing accidents, such as spillages in work area;
  • Keeping the laboratory and surrounding work areas clean;
  • Judicious use of antibiotics to prevent the formation of antibiotic-resistant strains;
  • Routine quality checks for contamination; and
  • Strategic use of frozen cell repositories are some of the steps to control contamination.
Aseptic Techniques: Cell Culture Basics

Placing adequate measures to reduce culture contamination can save the research industry from huge monetary losses as well as prevent health hazards in laboratory personnel who are routinely exposed to potent viruses (HIV, hepatitis B, Epstein Barr, herpes), bacteria and other microbes.

Further Reading

Last Updated: Oct 3, 2018

Deepthi Sathyajith

Written by

Deepthi Sathyajith

Deepthi spent much of her early career working as a post-doctoral researcher in the field of pharmacognosy. She began her career in pharmacovigilance, where she worked on many global projects with some of the world's leading pharmaceutical companies. Deepthi is now a consultant scientific writer for a large pharmaceutical company and occasionally works with News-Medical, applying her expertise to a wide range of life sciences subjects.


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