EZ-PCR™ Mycoplasma Detection Kit
Highly-sensitive PCR assay designed for the simple and fast detection of mycoplasma contamination in cell cultures. Includes ready-to-use reagents to perform 20 tests, each with internal control.
Easily detect contamination in cell cultures
The EZ-PCR™ Mycoplasma Detection Kit is a highly-sensitive PCR assay designed to test for the presence of over 90 species of Mycoplasma, Acholeplasma, and Spiroplasma in cell cultures with a detection limit of 10 CFU/mL. The kit contains a ready-to-use PCR reaction mix that includes highly-optimized mycoplasma-specific primers, dNTP mix, and Taq polymerase. Internal and positive controls are also included in order to test for false negatives and for simplistic result interpretation, respectively.
With a ready-to-use format and easy-to-follow protocol, samples can be prepared in as little as 10 minutes with accurate results seen in only a few hours, making it an ideal system for routine mycoplasma screening.
- Highly-sensitive PCR-based assay
- Simple protocol
- Prepare samples in less than 10 minutes
- Rapid and reliable results
- Ideal for routine testing
Figure 1. Gel electrophoresis results obtained following PCR reaction preparation and amplification. Eight total reactions, including six samples, one negative control, and one positive control were tested, each of which contain the internal control to rule out PCR inhibition (i.e. false negatives). As shown, Sample 4 produced a mycoplasma-positive band at 270bp in addition to the internal control band at 357bp, while Samples 1, 2, 3, 5, and 6 produced only internal control bands and are thus negative for mycoplasma. Image courtesy of WiCell Research Institute.
Figure 2. Images represent examples of mycoplasma that are capable of infecting all cell culture labs. Mycoplasma can be self-replicating and are the smallest known free-living prokaryote. Mycoplasma contamination can not be seen under standard microscopes. Regulatory guidance recommends that all products derived from mammalian cell culture be tested for the presence of mycoplasma.
The importance of routine testing
Mycoplasma are one of the most common, yet elusive, contaminants of mammalian cell cultures. As the smallest known free-living organism, mycoplasma are a pervasive, parasitic species of highly-infectious bacteria that are estimated to contaminate between 15-35% of all continuous cell cultures worldwide. While other mycoplasma detection methods are available, PCR-based assays have the highest sensitivity with minimal preparation time for early detection in a rapid and simple manner, when compared to other methods.
The earlier mycoplasma contamination is discovered, the simpler it is to treat. The easy-to-use and cost-efficient EZ-PCR™ Mycoplasma Detection Kit was designed for highly-sensitive routine screening and detection of mycoplasma and other closely related species. To avoid major contamination, testing should be carried out minimally every 2 weeks to 3 months, especially when shared incubator spaces are used, as well as prior to the incorporation of new cultures from outside sources.
Efficient and precise PCR-based testing for mycoplasma infection should also be conducted throughout the cell culture manufacturing process from inoculation through harvest, involving routine tests of raw materials, cell banks, and viral seed stocks.
|Intended use||Detection of mycoplasma contamination in cell cultures|
|Catalog Number(s)||20-700-20 (for 20 tests)|
|Kit Components||Reaction Mix (200μl)|
Buffer Solution (1.0ml)
Positive Template Control (20μl)
Internal control DNA template (20μl)
Internal control primers mix (100μl)
|Additional Components Required||Mineral Oil|
Agarose for gel electrophoresis
Sterile distilled water
|Storage||Store at -20°C <BR>Avoid repeated freeze/thaw cycles. When in use, always keep kit vials on ice.|
|Quality Statement||Manufactured by Biological Industries.|
This product is for Research Use Only and is not intended for therapeutic or diagnostic use.
References and Publications
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- R. M. Dulay, et al. Cytotoxicity of Gymnopilus purpureosquamulosus extracts on hematologic malignant cells through activation of the SAPK/JNK signaling pathway. (2021) PLoS ONE 16(5): e0252541. https://doi.org/10.1371/journal.pone.0252541
- A. Danilevsky, et al. Real-time selective sequencing using nanopores and deep learning. Research Square, 2021 https://doi.org/10.21203/rs.3.rs-540693/v1
- X. Gao, et. al. Generation of an iPSC line (GWCMCi002-A) from an X-linked Alport syndrome patient with a hemizygous splicing mutation (NM_000495.4, c. 1517-1 G > T) in the COL4A5 gene. Stem Cell Research, 2021, https://doi.org/10.1016/j.scr.2021.102388.
- R. Guo, et. al. Reprogramming of a human induced pluripotent stem cell line from one 48-year-old healthy male donor. Stem Cell Research, Vol. 53, 2021, https://doi.org/10.1016/j.scr.2021.102339.
- X. Wang, et. al. Derivation of induced pluripotent stem cells from one child suffering Potocki-Lupski syndrome. Stem Cell Research, Volume 53, 2021, https://doi.org/10.1016/j.scr.2021.102324.
- T.W. Hsu, et. al. Transplantation of 3D MSC/HUVEC spheroids with neuroprotective and proangiogenic potentials ameliorates ischemic stroke brain injury. Biomaterials, Vol. 272, 2021, https://doi.org/10.1016/j.biomaterials.2021.120765.
- T.Y. Chang, et. al. A novel histone deacetylase inhibitor MPT0L184 dysregulates cell-cycle checkpoints and initiates unscheduled mitotic signaling. Biomedicine & Pharmacotherapy, 2021, https://doi.org/10.1016/j.biopha.2021.111485.
- S. Kim, et. al. Inhibition of platelet‑derived growth factor receptor synergistically increases the pharmacological effect of tamoxifen in estrogen receptor α positive breast cancer. Oncology Letters, (2021). 21, 294. https://doi.org/10.3892/ol.2021.12555
- C. B. Lanauze, et. al. Colorectal cancer-associated Smad4 R361 hotspot mutations boost Wnt/β-catenin signaling through enhanced Smad4-LEF1 binding, Mol Cancer Res February 19 2021 DOI: 10.1158/1541-7786.MCR-20-0721
How many assays (or tests) are in each EZ-PCR kit?
Each kit has a Reaction Mix (200μl), Buffer Solution (1.0ml), Positive Template Control (20μl), Internal control DNA template (20μl), and Internal control primers mix (100μl) - enough reagents to conduct 20 tests.
How can I be sure that a faint positive band means my sample is mycoplasma positive?
To know for sure, a sample producing a faint positive band may be re-tested. If a faint band persists in the subsequent assay, the sample should be considered mycoplasma positive. If the faint band no longer appears in the subsequent assay, the sample is negative.
How definitive are the given gel electrophoresis parameters?
Running the 2% agarose gel at 100 volts for 75 minutes is a recommendation but may vary depending on the apparatus. If recommendations for specific gel electrophoresis setups are needed, one option is to prepare and run a pre-made 2% E Gel agarose gel (Thermo Fisher) according to protocol with the appropriate program on the E-Gel® iBase™ Power System (Thermo Fisher) to run for 26 minutes.
What happens if samples are collected for mycoplasma testing more than 48 hours after the last media exchange?
If cell culture media is older than 48 hours when collected, cellular byproducts can accumulate that can inhibit the PCR reaction. In this case, it is best to replace media and collect for PCR between 24 and 48 hours later. If a sample collected after the 48-hour timeframe must be used, DNA extraction is recommended prior to PCR amplification.