References
Strategies for
The Elimination of Biological Contamination from Laboratory Incubators
Mycoplasma contamination poses a significant challenge in cell culture laboratories, compromising research integrity and leading to inaccurate experimental results.
Estimates for the incidence of Mycoplasma contamination vary from 10% to 36% of cell lines used in laboratory procedures. Laboratory incubators, being a critical environment for cell growth, are prone to Mycoplasma contamination.
This paper explores effective strategies to eliminate Mycoplasma contamination from laboratory incubators, ensuring reliable and consistent cell culture experiments.
1
Cleaning and Decontamination Procedures
To eliminate biological contamination, thorough cleaning and decontamination protocols are essential. The following steps can be employed:
Regular Cleaning
Implement a stringent cleaning regimen for laboratory incubators, involving the removal of all cell culture vessels, shelves, and trays. Clean surfaces using appropriate disinfectants, such as 70% ethanol or other proven disinfectants, to effectively eliminate mycoplasma.
Disinfection
After cleaning, disinfect the incubator using (eg) heat treatment, Hydrogen Peroxide Vapor (VHP), Paracetic Acid (PAA), or Ultraviolet (UV) light. These methods are highly effective in eradicating mycoplasma and other contaminants.
Periodic Maintenance
Establish a maintenance schedule for incubators to ensure proper functioning and prevent biological contamination. This includes regular replacement of filters, inspection of seals, and calibration of temperature and humidity controls.
Isolation
Identify and segregate contaminated cell lines from healthy ones. Store contaminated cultures separately, preferably in sealed containers, to prevent cross-contamination.
Quarantine
Newly obtained or suspected cell lines should be quarantined for mycoplasma testing before integration into the laboratory. Maintain a dedicated space for quarantine and perform regular testing to ensure early detection and prompt elimination of contamination sources.
In some cases it may be possible to recover contaminated cultures using fluoroquinolone antibiotics. If there is no other source for uncontaminated cells, the time and expense may be justified.
2
Isolation and Quarantine
Preventing biological contamination requires isolating contaminated cell lines and practicing strict quarantine measures:
3
Regular Testing
Regular and rigorous testing protocols are indispensable in identifying Mycoplasma contamination and taking appropriate measures:
Testing Methods
Employ validated Mycoplasma detection methods, such as polymerase chain reaction (PCR) or DNA staining assays. These tests are highly sensitive and specific, allowing for accurate detection of mycoplasma contamination.
Frequency
Establish a periodic testing schedule to ensure the timely identification of Mycoplasma contamination. Regular testing, at least once a month, is recommended, especially for high-throughput laboratories with extensive cell culture activities.
External Testing
Periodically utilize external services or contract laboratories for Mycoplasma testing to obtain an independent and unbiased assessment of incubator and cell culture contamination.
By implementing robust cleaning and decontamination procedures, ensuring isolation and quarantine practices, and implementing regular mycoplasma testing, laboratories can significantly reduce the risk of contamination.
Maintaining a vigilant approach towards mycoplasma elimination is crucial for generating reliable and reproducible experimental results, thus promoting the advancement of scientific research.
Maintaining a vigilant approach towards mycoplasma elimination is crucial for generating reliable and reproducible experimental results, thus promoting the advancement of scientific research.
References
- Drexler H, Uphof CC (2002). In Vitro Cell. Biol. - Animal 38:79-85
- Olarerin-George AO, Hogenesch JB (2015). Nucleic Acids Research 43(5): 2535-2542
- Uphof CC, Denkman Sabine-A, Drexler HG (2012) J. Biomed. Biotechnol. 2012: 267678
- Chat-GPT 4, OpenAI
4
What makes tackling Mycoplasma contamination crucial?
Mycoplasma contamination represents a persistent challenge in laboratory incubators and can compromise the validity of cell culture experiments.
MycoFog's Testimonials
“So far, I have had a chance to use it twice on our incubators. We tested after the first biodecontamination cycle using settle plates and found no contamination where we were observing consistent contamination previously. Similar testing on a second incubator yielded similar results. The system is very easy to use, instructions are fine. I like your product.”
- Donald Pijak,
Manager, Research Facility Operations, University of Pennsylvania
“We were looking for new ways to decontaminate our incubators and hypoxic workstation. The MycoFog™ proved to be an effective means of keeping the interiors sterile and free of contaminants. The decontamination cycle takes only 180 minutes and leaves no H2O2 residue after the procedure. The MycoFog™ is light-weight and easy to set up, generating a fog that reaches all surfaces inside the incubator, and is proving to be an extremely valuable addition to our contamination control practices.”
- Dr. William Willmore, Professor of Biochemistry, Carleton University, Ottawa
"We have now used the MycoFog™ system in two of our eight incubators and have thus far found it to be very easy to figure out and implement. In both instances, adhering to the biological indicator test guidelines, we've observed consistent results aligning with our expectations.
We have had persistent contaminations in the past for these two incubators. There has been no contamination observed in either incubator since first using the system 7 weeks ago. Overall, it is a good system that is easy enough to integrate into lab protocols and workflows!”
- Deborah Schwarz , CEO , Advanced Cellular Dynamics, Seattle, WA
validation
The definitive validation of disinfection processes is the use of spores of the bacteria Geobacillus stearothermophilus. These spores are extremely hardy and are used to validate biodecontamination and disinfection processes in number of applications.
7.8 BIO tests the MycoFog™ biodecontamination process by placing the biological indicator (BI) (from Mesa Labs or Terragene) which are stainless steel discs impregnated with at least 1 x 106 Geobacillus stearothermophilus (G.s.) spores, into the incubator. The device is sealed so that it is required that the sterilant (nebulized hydrogen peroxide, nHP) penetrate a membrane to come into contact with the spores, killing them.
7.8 BIO tests the MycoFog™ biodecontamination process by placing the biological indicator (BI) (from Mesa Labs or Terragene) which are stainless steel discs impregnated with at least 1 x 106 Geobacillus stearothermophilus (G.s.) spores, into the incubator. The device is sealed so that it is required that the sterilant (nebulized hydrogen peroxide, nHP) penetrate a membrane to come into contact with the spores, killing them.
Locations in the incubator where the G.s. biological indicator devices (BI’s) are placed for a validation run.
The BI’s are placed in the appropriate position in the incubator and exposed to a full biodecontamination cycle, processed, and then incubated for 7 days.
Results from one validation run of Geobacillus stearothermophilus test devices described above. The control device is outside of the incubator chamber while the test devices are exposed to a complete biodecontamination cycle in the incubator. The media color changes to yellow when the spores grow, negative growth is indicated by the purple color. These results indicate successful biodecontamination of the incubator.