Cell lines are important research tools, and contamination with microorganisms is one of the most common concerns. The cell lines need to be authentic to provide the most reliable results. This is why many grants require researchers to test cell lines for mycoplasma contamination.

Why Contamination Is a Problem

Mycoplasmas are the smallest, simplest form of self-replicating bacteria. As surface parasites, they can be found in the human urogenital and respiratory tracts. Since the bacteria don't have cell walls, many common antibiotics have no effect in health treatments.

In research, cell culture contamination with mycoplasmas can cause a myriad of problems, interfering with nearly every aspect of cell culture research. In many cases, it can result in the degeneration or total loss of the cultures. It can lead to changes in cell membrane composition, DNA synthesis, protein levels, reproductive characteristics and cellular metabolism. Contamination can also influence cellular transformation, virus reproduction and cell fusion.

Detecting and Eliminating Mycoplasmas

The wide ranging effects of mycoplasma contamination are cause for many grant requirements to include testing for the microorganisms. Various methods have been developed to detect mycoplasmas in cell lines. Ideally, the methods that researchers use should be very specific and sensitive as well as fast, efficient, cost effective and simple.

Researchers may use elimination methods to eradicate the mycoplasmas that they find. However, it's recommended that they only use this as a last resort to avoid spreading the contamination. Most of the developed methods are also unreliable.

Preventing Mycoplasma Contamination

The Department of Human and Animal Cell Cultures in Braunschweig, Germany, proposes changes at three levels to prevent mycoplasmas from contaminating cell lines. Although the suggested measures won't automatically prevent contamination, they'll greatly reduce the incidence.

The Facility

The cell culture facility itself should have a vertical laminar-flow biohazard hood. Culturists much disinfect every work surface before and after culture manipulations. They even need to disinfect the devices that enter the hood. Although mycoplasmas can survive in a dried state for an extended period, they're very sensitive to most disinfectants. Other facility recommendations include

• not keeping animals and cell cultures in the same room,
• barring entry to unauthorized persons, and
• properly disposing cell culture materials through central sterilization.

Testing Procedures

Facilities must establish effective and routine procedures to test for mycoplasmas. However, routine screening programs should include all forms of microbial contamination. The department recommends polymerase chain reaction analysis. It also suggests that facilities purchase media, sera and supplements from reputable suppliers that perform adequate testing. Other procedure recommendations include

• putting all incoming cultures into quarantine until culturists can verify their contamination status,
• separating negative cultures from infected cultures by place and time of handling, and
• performing screenings at the time of arrival and monthly.

Culturists need to adhere to strict sterile techniques, which is a basic element in preventing contamination. They should only handle one cell line at a time, remove all jewelry and tie back long hair. Other technique recommendations include

• keeping written laboratory records for each cell culture,
• wearing protective clothing,
• not applying cosmetics, eating, drinking and smoking in the lab, and
• washing and disinfecting hands before and after work.

By using efficient methods to detect and prevent mycoplasma contamination, researchers can avoid losing cultures. They can also expect more reliable results from their experiments and may have a greater chance of earning grant funds.