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Product Description
MycoSHENTEK® Mycoplasma DNA Detection Kit (2G) is a qualitative qPCR-based assay for detection of Mycoplasma, Spiroplasma and Acholeplasma contamination in master cell banks, working cell banks, virus seed stocks and cell culture–derived products. The kit is validated according to USP <63>, EP 2.6.7 and JP XVIII, with a stated limit of detection of 10 CFU/mL. The assay detects >200 species with high specificity and includes Internal Control (IC, 2G) to monitor PCR inhibition and optionally to evaluate extraction efficiency when added prior to extraction. The kit is supplied for 50 reactions and is intended for research use only (RUO).
Technical Specifications
| Parameter | Details |
|---|---|
| Assay type | Qualitative real-time PCR (qPCR) assay |
| Limit of detection (LOD) | 10 CFU/mL (validated) |
| Validation / Standards | Validated according to USP <63>, EP 2.6.7 and JP XVIII |
| Species coverage | Detects >200 species of Mycoplasma, Spiroplasma and Acholeplasma |
| Controls included | Internal Control (IC) (2G), Positive Control (PC) (2G) |
| Reaction format & volume | 30 μL total reaction volume: 10 μL qPCR MIX + 20 μL sample (per well) |
| qPCR Mix composition (per reaction) | My qPCR Reaction Buffer (2G) 8 μL; My Primer & Probe MIX (2G) 1.5 μL; Internal Control (IC) (2G) 0.5 μL (total 10 μL) |
| Thermal cycling conditions | UNG 25°C 10:00 (1); Activation 95°C 10:00 (1); 45 cycles of: Denaturation 95°C 00:15, Annealing 62°C 00:30, Extension 72°C 01:30 (fluorescence read during extension) |
| Detection channels / dyes | Target: FAM; Internal Control: VIC; ROX optional as passive reference |
| Instrument compatibility | SHENTEK-96S, Bio-Rad CFX96, Applied Biosystems 7500, Roche LightCycler 480 and other real-time PCR systems |
| Intended sample types / matrices | Master cell bank, working cell bank, virus seed stock, cell culture-derived products and similar matrices (validated across a variety of matrices) |
| Total reactions per kit | 50 reactions |
| Storage & stability | -20°C; kit components stable up to 24 months (check label expiration date) |
| Reconstitution instructions (controls) | Reconstitute lyophilized IC (2G) in 600 μL DNA Dilution Buffer; reconstitute lyophilized PC (2G) in 500 μL DNA Dilution Buffer |
| Acceptance criteria (Ct thresholds) | NTC: FAM Ct ≥ 40 or no amplification; VIC Ct < 35; PC: FAM Ct < 35; Unknown positive: FAM Ct < 40 (≥1 well) with normal amplification; If VIC inhibited, repeat extraction or remove inhibitors |
Features
- High analytical sensitivity: Validated LOD of 10 CFU/mL allowing detection of low-level mycoplasma contamination
- Broad species coverage: Designed to detect more than 200 Mycoplasma, Spiroplasma and Acholeplasma species with high specificity
- Internal control for inhibition and extraction monitoring: Internal Control (IC, 2G) can be added to PCR or to the sample prior to extraction to monitor PCR inhibition and evaluate extraction efficiency
- Validated to pharmacopeia standards: Validated according to USP <63>, EP 2.6.7 and JP XVIII
- Ready-to-use primer/probe mix: Provided My Primer & Probe MIX (2G) simplifies assay setup; lyophilized positive and internal controls included
- Platform compatibility: Compatible with common real-time PCR instruments (SHENTEK-96S, CFX96, 7500, LightCycler 480, etc.)
Applications
- Qualitative mycoplasma testing of master cell banks and working cell banks
- Screening of virus seed stocks for mycoplasma contamination
- Testing of cell culture–derived products and biologics for mycoplasma
- Routine cell culture contamination monitoring and QC in R&D and production (research use only)
Kit Contents
Internal Control (IC) (2G) (Part No. NNA035) - lyophilized-20°C (reconstitute in 600 μL DNA Dilution Buffer)
Positive Control (PC) (2G) (Part No. NNA039) - lyophilized-20°C (reconstitute in 500 μL DNA Dilution Buffer)
My qPCR Reaction Buffer (2G) (Part No. NNB004) 400 μL × 1 tube-20°C, protect from light
My Primer & Probe MIX (2G) (Part No. NNC065) 75 μL × 1 tube-20°C
DNA Dilution Buffer (DDB) (Part No. NND001) 1.5 mL × 2 tubes-20°C
User Guide / Instructions for useRoom temperature (follow handling instructions)
* Total: 50 Reactions | Method: qPCR
Attention
• For research use only (RUO). Not for use in diagnostic procedures.
• Read the Material Safety Data Sheets (MSDS) and follow handling instructions.
• Wear appropriate PPE: protective eyewear, mask, clothing and gloves.
• Work in a clean area: irradiate work surfaces, pipettes and tubes with UV for 30 minutes and disinfect with 75% ethanol before use.
• Thaw kit components at 2–8°C or on ice; vortex and centrifuge briefly before use.
• Protect My qPCR Reaction Buffer from light during storage and use.
• If Internal Control (IC) was added during extraction, substitute equal volume of DNA Dilution Buffer in qPCR mix instead of IC.
• Follow recommended plate layout and include controls (PC, NTC, NCS) in duplicate.
• If VIC (IC) signal is inhibited (Ct ≥ threshold), repeat extraction or remove inhibitors and repeat assay.
• Dispose of biological and chemical waste according to local regulations.
Quality Management & Certifications
Quality System
Download QMSQMS (ISO, GMP)
Download CertificateQuality Advantages
View ReportQuality Control Process
Download ProcessTechnical Resources & Downloads
Frequently Asked Questions (FAQ)
Q1: What is the limit of detection (LOD) of the kit?
The kit is validated with a limit of detection of 10 CFU/mL.
Q2: Can the Internal Control (IC) be added before extraction?
Yes. IC (2G) may be added to the sample prior to extraction to evaluate extraction efficiency; if IC is added during extraction, omit IC when preparing the qPCR mix and add equal volume of DNA Dilution Buffer instead.
Q3: Which qPCR instruments are compatible with this kit?
The kit is compatible with common real-time PCR platforms including SHENTEK-96S, Bio-Rad CFX96, Applied Biosystems 7500 and Roche LightCycler 480. Use instrument software to set FAM for target and VIC for IC.
Q4: How are results interpreted (Ct thresholds)?
Acceptance criteria: Positive control (PC) FAM Ct < 35; NTC FAM Ct ≥ 40 or no amplification; Unknown sample: FAM Ct < 40 (at least one well) with normal amplification = Positive. If IC (VIC) is inhibited (VIC Ct ≥ threshold or absent), repeat extraction or remove inhibitors and retest.
Research Use Only
✔
Research Use Only (RUO) – This product is intended for laboratory research purposes only and not for clinical or regulatory diagnostic use.
✔
Not intended for use in USDA or FDA regulated diagnostic testing or official compliance testing.
When can I expect my order to ship?
Most orders are filled and shipped within 2-3 business days from the time they are received.
Our standard shipping usually take 2-5 days.
We also provide express shippping for time-sensitive deliveries.
Email contact@biofargo.com if you have any requirements.
