Mycoplasma contamination remains a major problem for researchers using mammalian cell culture. The Myco Scan Kit from Applied Biosystems® can be used to do rapid and reliable Mycoplasma contamination using Real-Time PCR. Through intensive bioinformatics and highly optimized multiplexed primer design, the assay allows for highly specific and comprehensive Mycoplasma species detection and prevents the detection of other closely related species. Providing results in less than five hours, the kit is a critical tool in testing the authenticity of valuable cell lines used in basic research.

Mycoplasma contamination, often referred to as the “stealth invader" or “silent killer" of high-value cell lines, is common in many laboratories and is a major problem for scientists whose research involves cultured mammalian cells. Because Mycoplasma infection can affect virtually every cell parameter, many studies have underlined the need for routine screening of research cell lines to help ensure that the results observed are not compromised by contamination.

• Helps ensure mycoplasma-free cell lines
• Rapid time-to-results—under five hours
• Detection of greater than 90 species
• Demonstrated sensitivity and specificity
• Efficient sample preparation allows high DNA yields

Routine Testing As Part of Good Cell Culture Practice
The European Collection of Cell Cultures (ECACC) states, “The effects of Mycoplasma infection are more insidious than those of bacteria and fungi. Mycoplasma contamination is difficult to detect, requiring the use of specialist techniques. These tests should be used as part of a routine and regular quality control screening ´procedure." [1] And an open letter to the National Institutes of Health regarding misidentification and cross-contamination of cells [2], recommends that laboratories adhere to Good Cell Culture Practice (GCCP) guidelines [3] that include a quality assurance regimen requiring Mycoplasma testing when new cell lines are introduced into a laboratory, followed by routine screening to help ensure that they remain free of Mycoplasma contamination [4].

Rapid Time-to-Results—Under 5 Hours
The Myco Scan Kit has an easy workflow that can deliver results in less than 5 hours (Figure 1), which helps to facilitate routine screening of cell lines.
Key features include:
• Rapid and high-recovery sample preparation protocol
• Sensitivity levels of less than 10 CFU⁄mL
• Detection of greater than 90 species
• Single-step detection
• Load-and-run, walk-away automation
• No gel electrophoresis
• Minimal demands on infrastructure and space

Detection of >90 Mycoplasma Species
For comprehensive detection of Mycoplasmas, an assay must have the ability to detect all known Mycoplasma species, not just the most common species. Intensive bioinformatics analysis was utilized to design amplification primers and reaction conditions to enable comprehensive detection of Mycoplasma, Acholeplasma, and Spiroplasma species (Table 1).

Demonstrated Sensitivity and Specificity
The Myco Scan Kit has a limit of detection of fewer than 10 copies per reaction (Figure 2). It is specific for Mycoplasmas and does not detect DNA purified from related and unrelated species and organisms.

High DNA Recovery Sample Preparation
The Myco Scan Kit is optimized for use with the PrepSEQ™ 1-2-3 Mycoplasma Sample Preparation Kit, which uses magnetic bead–based separation technology to extract mycoplasma DNA from mammalian cell lines. The PrepSEQ™ 1-2-3 Kit can be used to extract Mycoplasma genomic DNA from research cell lines, stem cell lines, and cell- and tissue-therapy samples. In the PrepSEQ™ 1-2-3 Kit protocol, a rapid procedure recovers DNA from total lysate of the test sample.