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| Product Name | Mycoplasma PCR Detection Kit (Nested PCR, 2G) |
| Catalog No. | RESK-0191 |
| Description | The Mycoplasma PCR Detection Kit (Nested PCR, 2G) is based on the nested PCR method for the detection of mycoplasma infection in various biological materials (such as cell cultures, laboratory animal secretions, and animal serum). The kit uses PCR technology with two sets of primers designed against the conserved region of the mycoplasma 16S-23S rRNA sequence, performing two rounds of PCR amplification; mycoplasma contamination is judged based on the second-round amplification product. It can detect a total of 34 mycoplasma species, including M. orale, M. pneumoniae, M. hyorhinis, M. arginini, M. gallisepticum, M. synoviae, M. fermentans, Spiroplasma citri, and M. salivarium, with a sensitivity of up to 2.5 copies/uL; mycoplasma standard strains at 10 CFU/mL can also be amplified normally with visible bands. Featuring strong specificity, high sensitivity, and convenient and rapid operation, the kit can be used directly for the detection of cell culture supernatants. Compared with mycoplasma detection kits based on conventional PCR, this product avoids the generation of nonspecific products caused by nonspecific pairing between primers and template, and can sensitively detect trace amounts of mycoplasma DNA in samples, avoiding false negatives. |
| Intended Use | For the detection of mycoplasma infection in various biological materials, such as cell cultures, laboratory animal secretions, and animal serum; suitable for direct testing of cell culture supernatants. |
| Principle / Technology | Nested PCR with two sets of primers designed against the conserved region of the mycoplasma 16S-23S rRNA sequence. Two rounds of PCR amplification are performed, and the result is interpreted based on the second-round amplification product. Compared with conventional PCR, nested PCR avoids nonspecific products caused by nonspecific primer-template pairing and reduces false negatives by sensitively detecting trace amounts of mycoplasma DNA. |
| Detection Method | Two rounds of nested PCR amplification followed by agarose gel electrophoresis analysis; results are judged based on the second-round amplification product. |
| Sample Type | Cell culture supernatants (can be tested directly), laboratory animal secretions, animal serum, and other biological materials. |
| Performance Range / Specifications | Broad detection coverage: the optimized nested PCR technology detects up to 34 mycoplasma species. Strong specificity: primers and probes designed against the 16S-23S rRNA region show no cross-reactivity with related species such as Clostridium and Lactobacillus. High sensitivity: the limit of detection can be as low as 2.5 copies/uL. Simple operation: the total time for sample preparation and detection can be as short as 2 h. High safety: the positive control (PC) included in the kit is non-infectious, completely eliminating contamination risk. All 8 mycoplasma strains required by international pharmacopoeias were detected at the pharmacopoeia-required sensitivity of 10 CFU/mL: M. fermentans, Spiroplasma citri, M. hyorhinis, M. pneumoniae, M. orale, M. synoviae, M. arginini, and M. salivarium (M. gallisepticum and Acholeplasma laidlawii were not tested due to insufficient samples). |
| Sensitivity / LOD | First-round PCR amplification: 1 x 10^3 copies/uL. Second-round PCR amplification: 2.5 copies/uL. |
| Specificity | No cross-reactivity with related species such as Clostridium and Lactobacillus. Matrix interference was validated with 14 common sample matrices (MEM (1x), MCDB 131, F-12, IMDM, BME, McCoy's 5A medium, DMEM, opti-MEM, RPMI medium, L-15 Leibovitz medium, Ham's F-12 nutrient mixture, RPMI medium modified, Medium 199/EBSS, and DMEM/low glucose), the kit positive control and its spike (2.5 copies/uL positive control), and the no-template control; no mycoplasma was detected in any of the 14 matrices. |
| Reaction Conditions / Protocol | The whole experiment consists of 2 rounds of PCR amplification, and the test result is based on the second round. The total time for sample preparation and detection can be as short as 2 h. |
| Components / Formulation | Includes a non-infectious positive control (PC), completely eliminating contamination risk. |
| Storage Conditions | Store at -25 to -15 °C. If the whole kit will not be used for an extended period, store it protected from light. Components not used for a long time may be stored frozen at -85 to -65 °C. |
| Shelf Life | 1 year. |
| Package Specifications | 10 assays / 20 assays |
| Shipping Conditions | Shipped on dry ice. |
| Regulatory / Compliance | For research use only. |
| Application Notes / Precautions | Compared with the basic conventional-PCR mycoplasma detection kit, this upgraded 2G nested PCR kit offers stronger specificity, higher sensitivity, and a broader range of detectable mycoplasma species. |
For research use only, not for clinical use.
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