Detecting and Managing Mycoplasma and Nanobacteria Contamination

Cell contamination is the most common and challenging problem in cell culture. When abnormal cell morphology is detected, it is important to promptly investigate and handle the contamination, otherwise it will have a significant impact on our experiments.

Among various types of contamination, we found that the most troublesome contamination is "black spots". Therefore, we will introduce the possible types of contamination when cells show black spots: Mycoplasma contamination and Nanobacteria contamination.

Mycoplasma Contamination

Mycoplasma can almost affect any data on cell growth, cell metabolism and cell research. Moreover, conventional antibiotics cannot resist Mycoplasma because they lack cell walls, and their small size renders them unable to be filtered by standard filters.

Figure 1 Mycoplasma Contamination

Common manifestations of Mycoplasma contamination:

1. The cell state deteriorates and growth slows down, and there may be small black spots observed by the microscope, but the culture medium is not cloudy;

2. After cell passage, there are black spots, cell vacuolization, or many cells that resemble apoptosis or necrosis, ultimately leading to cell floating and complete death.

Nanobacteria Contamination

Nanobacteria and their decomposition complexes are common cellular contaminants that coexist with cells, and they could passaged through cell passages. Antibiotics are usually ineffective against them. Nanobacteria compete with cells for growth, which is detrimental to cell growth and can lead to cell death in severe cases.

Figure 2 Nanobacteria Contamination

Common manifestations of Nanobacteria contamination

• The culture medium is not turbid, but when observing cells by the microscope, there are many "small black spots" around the cells and in the culture medium, and as the culture time prolongs, the number of "small black spots" gradually increases. It’s hard to remove them by changing the culture medium or washing the cells.
• Cells consume nutrients quickly and require refresh culture media frequently.
• The cell growth is slow, the cell condition is poor, the vacuolation is serious, and the cell morphology may even change.

Handling method

• If the cell condition is severely compromised, it is recommended to discard it directly and clean the incubator from the inside out;
• If the cell condition is normal, then in the case of determining that there is no effect on the gene and protein expression of the cell, the Mycoplasma removal reagent or Nanobacteria removal reagent can be used accordingly, and culture for 2-3 generations can turn negative.

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