Mycoplasma Detection: Bioluminescent-based and PCR-based
It was estimated that mammalian cells in up to 60% of the published papers were already contaminated by mycoplasma. Mycoplasma contamination in cells can alter many cellular parameters, leading to unreliable experimental results, misleading data interpreting and potentially unsafe biological products. The big issue is that many labs do not have a routine cell culture screening schedule to monitor mycoplasma contamination in tissue culture labs. We thus developed both PCR-based and bioluminescent-based assays, to rapidly and accurately detect mycoplasma contamination. The flowchart of the bioluminescent-based assay was showed in Fig.1.
The principle of this bioluminescent-based assay is based on the observation, which mycoplasma can efficiently and rapidly breakdown luciferase enzyme. The presence of mycoplasma contamination in cell culture would decrease the luciferase activity while the luciferase activity in the mycoplasma-free cell culture supernatants would keep no change. Therefore, luciferase activity was inversely proportional to the extent of mycoplasma contamination.
Fig.1. The flowchart of Bioluminescent-based Mycoplasma Detection. 1. Overnight cell culture was prepared. 2. Change cell culture media to 1x Assay Media which contains luciferase. 3. Collect supernatants of cell culture at the 24hrs, 48hrs and 72 hrs time points, respectively. 4. Measure luciferase in the supernatants at the luminescent reader.