MycAway™ 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. Validation Report Specifications Sample Type media, cells, raw materials; Biopharmaceutical Purification Sample, Bulk Drug Substance Sample, Cell Cultures. Detect method qPCR method(Taqman fluorescent) Detect Time 95% detection rate. Figure 3. Limit of detection (LOD) test. Sample FAM Ct CY5 Ct Interpretation Mycoplasma arginini 10 CFU/mL 33.36 26.96 Positive 33.28 26.96 Positive 33.60 26.95 Positive 33.57 27.00 Positive 33.81 27.26 Positive 33.48 27.12 Positive 33.77 27.02 Positive 32.20 26.88 Positive NCS / 27.22 valid NCS / 27.32 valid NTC / 27.26 valid NTC / 26.92 valid Table 2. Example detection results for Mycoplasma arginini 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. 4. Performance Validation Summary Performance Validation parameter Kit validation result Limit of detection LOD for mycoplasma strains 10 pharmacopoeia-required mycoplasma strains at 10 CFU/mL, each tested 24 times; detection rate ≥ 95% Specificity Sample matrix interference 9 sample matrices and DNA dilution buffer tested — no mycoplasma detected Specificity Cross-reactivity 14 strains and 6 commonly used biopharmaceutical engineered cell lines tested — no mycoplasma detected Robustness Freeze-thaw stability Withstands 20 repeated freeze-thaw cycles without affecting kit performance Robustness Heat-accelerated stability 14 days at 37°C and 30 days at 4°C without affecting kit performance Robustness Instrument compatibility Compatible with ABI 7500, ABI QuantStudio™ 5, Bio-Rad CFX96, Roche LightCycler 480 Coverage Mycoplasma DNA coverage Database comparison based on mycoplasma 16S rRNA sequences; covers 183 species of the class Mollicutes Table 3. Performance Validation Content and Conclusion of the Mycoplasma qPCR Detection Kit Documents: Safety Data Sheet 40619_MSDS_HB250814_EN.PDF Manuals 40619_Manual_Ver.EN20250814.pdf Related Blogs: Co-publish papers: Establishment of Nucleic Acid Amplification Technology for the Detection of Mycoplasma in Biological Products. Molecules 2026, 31(11), 1794. From Research to Regulatory Practice: Yeasen and NIFDC Advance NAT-Based Mycoplasma Testing for Biologics 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 development
