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Description
MycAwayTM Mycoplasma Real-time qPCR Detection Kit (2G) is a product which can qualitative detect the mycoplasma contamination in the raw materials, cell bank, virus seeds, virual or cell harvesting solution and cells used in clinical treatment, etc. The kit uses the Taqman fluorescent probe (which included FAM and CY5) and the Multiple polymerase chain reaction (PCR) tools to detect the target and internal control separately. It covered over 183 species of the Mollicutes DNA, the specificity, detection limit and robustness of this kit are validated according to EP2.6.7 which with high sensitivity, specificity, efficiency and safety. The detection limit is equals to and below 10 CFU/mL.
This product can be used in combination with Magnetic Residual DNA Sample Preparation Kit (Cat#18461ES/18469) which using the manual extracted method for the nucleic acid extraction. (Please note, the kits which included Cat#18461ES and Cat#40619ES are full validated, please contact our technical support for detailed validation information). After the samples are pre-treated to remove the interference impurities and obtain purified nucleic acid, then a qPCR reaction perform by the Real Time PCR amplifier and the fluorescence signal of the probe will be collected and analyzed.
Features
Broad Detection Range: Optimized TaqMan probes detect up to 183 mycoplasma species.
Fast & Convenient: Sample prep and testing completed in under 3 hours, versus 28 days for culture(Alternative to the standard 28-day culture test).
High Sensitivity: Detects as low as 10 CFU/mL, making it a reliable alternative to culture methods.
High Specificity: 16S rRNA-based primers and probes avoid cross-reactivity with related species such as Clostridium and Sesamum.
Safe to Use: Non-infectious positive control eliminates contamination risk.
Strong Anti-Interference: Internal control (IC) detects sample inhibition or reaction errors, reducing false negatives.
Regulatory Compliance: Validated according to the requirements of EP2.6.7, JP G3 and USP 63 pharmacopoeia, in line with the standards of international authorities.
Specifications
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Sample Type |
media, cells, raw materials; Biopharmaceutical Purification Sample, Bulk Drug Substance Sample, Cell Cultures. |
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Detect method |
qPCR method(Taqman fluorescent) |
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Detect Time |
<4 hours |
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Fluorescent probe |
FAM(Target channel)；VIC(Internal channel) |
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Pretreatment kit |
Compatible with magnetic bead-based sample pretreatment kits(Cat#18461/18469) |
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Applicable Models |
Thermo: ABI7500, ABI QuantStudio™ 5; Roche: LC 480; Bio-Rad：CFX96 Optic Module |
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Detection Limit(LOD) |
10 CFU/mL |
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Covered mycoplasma |
183 mycoplasma species |
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Validation |
Validated according to EP <2.6.7> |
Components
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Components No. |
Name |
40619ES25 |
40619ES60 |
|
40619-A |
2×MyqPCR Reaction Buffer |
375 μL |
1.5 mL |
|
40619-B |
MyPrimer & Probe Mix |
100 μL |
400 μL |
|
40619-C* |
Internal Control (IC) |
25 μL |
100 μL |
|
40619-D** |
Positive Control (PCS) |
250 μL |
1 mL |
|
40619-E*** |
DNA Dilution buffer |
500 μL |
2×1 mL |
|
40619-F**** |
Ultrapure water |
500 μL |
2×1 mL |
[Note]: *IC: Internal control.
**PCS: Positive control solution，the concentration is 1,000 copies/µL.
***DNA Dilution buffer: used for IC dilution and the template of NTC and NCS.
****Ultrapure water: used for the preparation of qPCR Mix.
Storage
This product should be stored at -25~-15℃ for 2 years.
*Upon receipt of the kit, please check whether all components are complete and immediately store them in -25~-15℃ condition if not perform the assay immediately. Please note 40619-B should be stored away from light.
Application
Mycoplasma Detection; Mycoplasma Detection for BioProduction process media, cells, raw materials.
Figures
1. Workflow
Figure 1. Workflow of mycoplasma detection using the Mycoplasma Real-time Quantitative PCR Detection Kit
2. Specificity
Figure 2. Specificity validation.
(A). Nine common sample matrices and the kit’s DNA dilution buffer showed no mycoplasma detection in the target channel (FAM, blue). Normal FAM signal was observed only when Mycoplasma fermentans was added to a sample matrix.
(B). DNA from 14 non-Mycoplasmatales strains and 6 commonly used biopharmaceutical cell lines showed no target channel amplification. In all cases, the internal control channel (Cy5, green) amplified normally.
3. Limit of Detection (LOD): >95% detection rate.
Figure 3. Limit of detection (LOD) test.
Ten mycoplasma standard strains (10 CFU/mL), sourced from German MB, were tested following kit instructions for nucleic acid extraction(Cat#18461) and detection kit(Cat#40619). Mycoplasma samples were tested three times with 8 replicates each. Each run included NCS (negative control solution) and NTC (no template control).
When controls passed, at least 23 out of 24 replicates tested positive for each strain.
Documents:
Related blog：
MycAway Mycoplasma Real-time qPCR Detection Kit (2G) Validation Report
Fast, and Accurate Mycoplasma qPCR Kit
Concept of Mycoplasma and the Impact of Contamination
Mycoplasma qPCR detection and method validation contribute to the advancement of cell and gene therapy technology developmentMycoplasma contamination is one of the most prevalent and insidious threats in cell culture, with estimates suggesting 15–35% of cell lines are contaminated at any given time. Unlike bacterial contamination, mycoplasma does not cause visible turbidity. Contaminated cells exhibit altered metabolism, gene expression, proliferation rates, cytokine secretion, and receptor signaling — invalidating experimental results. Regular testing is mandatory before publication, biobanking, or use in drug discovery assays.
This kit targets the most common cell culture contaminants, including Mycoplasma hyorhinis, M. orale, M. fermentans, M. arginini, M. hominis, Acholeplasma laidlawii, and others. Universal primer or probe designs used in PCR/qPCR-based kits detect conserved mycoplasma-specific sequences and may detect up to 70+ species. Refer to the species coverage table in the product datasheet for validated strains.
Best practices recommend testing all incoming cell stocks, after every 2–4 weeks in continuous culture, before cryopreservation, before experiments generating data for publication, and after any culture manipulation (transduction, transfection, co-culture). For GMP and biopharmaceutical environments, testing frequency and documentation must comply with regulatory agency guidelines (ICH Q5A/EMA/FDA).
Protocol requirements vary by kit format. MycAway colorimetric kits use culture supernatant directly (no extraction required), providing results in approximately 2 hours. PCR/qPCR-based kits (GMyc-PCR, qPCR variants) may require a brief extraction or heat lysis step for optimal sensitivity. The qPCR format provides quantitative data and is preferred for regulatory submissions. Refer to the specific product datasheet for the validated input protocol.
Yeasen Biotechnology offers flexible customization options for many of its assay kits and detection reagents, including custom lot sizes, bulk ordering, and application-specific formulation adjustments. Volume pricing, custom packaging, and kit bundling may be available depending on the product and intended workflow. A Certificate of Analysis (CoA) and lot-specific QC data are provided with every order. For inquiries regarding large-volume orders, custom configurations, or integration into automated workflows, please contact the BioHippo team for a tailored quotation.
