e-Myco™ VALiD-Q Mycoplasma qPCR Detection Kit
Fast and simple real-time PCR detection kit that replaces the direct method of mycoplasma negative testing
• Demonstrated the performance to satisfy the negative test of Mycoplasm based on nucleic acid amplification method of domestic, US and European Pharmacopoeia
• Suitable for process inspection before, during, and after manufacture of pharmaceuticals in regenerative medicine fields such as cell therapy and contamination monitoring
• High sensitivity (10 cfu / ml level) as an alternative to the direct method
• High specificity that does not detect other related microorganisms (including 3 strains related to domestic negative test validation)
• Closed pipe can be checked to prevent contamination by the result and use real-time PCR method to check the result within up to 3 hours of use
• Easy handling, minimizing deviation or contamination between experimenters
Mycoplasma is a serious contaminant commonly observed in cell culture. It is known that about 30-87% of cell cultures are infected with Mycoplasma. This can happen during the course of experiments (through humans), sometimes through animal-derived products (such as serum). Mycoplasma, unlike other germs, has no cell wall and can pass through the pore of the filter and can grow without being affected by antibiotics that inhibit cell wall synthesis. In addition, it is difficult to see under a microscope, and it affects the growth of contaminated cells but does not kill them. Therefore, culture is continuously performed without being aware of contamination by Mycoplasma.
e-Myco ™ VALiD-Q Mycoplasma qPCR Detection Kit is a product that amplifies a specific gene of Mycoplasma through PCR and detects Mycoplasma in real time using the fluorescence of the amplification product-specific probe. The Mycoplasma assay (culture method), which is currently used for quality control of cell therapy products, has a very long test period and is very short effective because it is made for testing of cell substrates used for the production of vaccines and monoclonal antibody drugs, which are produced in large quantities. Due to various limitations (sample volume, sample volume and media volume), it is difficult to apply existing test methods as they are during final quality control (shipping test).
This product is an e-Myco ™ VALiD-Q Mycoplasma qPCR Detection Kit, which is a successor to the existing e-Myco ™ VALiD Mycoplasma PCR Detection Kit. This product can be detected specifically. This product targets 16S Ribosomal RNA region of Mycoplasma and amplifies Mycoplasma target gene with high specificity and sensitivity, and Probe with complementary sequence to specific nucleic acid sequence inside target gene amplification product has a fluorescent color A fluorescent reporter dye is labeled and a quencher is labeled at the 3 'end, which is decomposed by DNA polymerase during amplification of the target gene. You can do it. This product has higher specificity than the PCR method because it is affected by the specificity of the probe along with the primer. In addition, it includes an internal positive control, which is designed to detect the IPC signal along with the target signal, ensuring the reliability of the analysis.
[How to use e-Myco ™ VALiD-Q Mycoplasma qPCR Detection Kit]
01 Monitoring Mycoplasma Contamination Before, During and After Cell Production
02 Contamination Management of Cell Lines
03 TA cloning for Mycoplasma detection in specimens and cell lines
|2X qPCR Master Mix Solution||-25 ~ -15℃|| 280 μl x 2 tubes|
(10 μl / test × 50 tests)
|Mycoplasma Detection Solution||-25 ~ -15℃||140 μl x 2 tubes|
|Positive Control (External PC)||-25 ~ -15℃||50 μl x 3 tubes|
|DNase/RNase Free Water||-||1 ml x 1 tube|
In order to confirm the detection specificity of Mycoplasma of this product, amplification was performed on 6 bacterial DNAs related to Mycoplasma and positive control M. hyorhinis, including 3 bacteria from the KFDA. MAM hyorhinis was able to confirm that the target signal FAM signal was amplified, and the positive control and 6 bacterial DNAs detected the internal control signal HEX signal. This confirms that e-Myco ™ VALiD-Q Mycoplasma qPCR Detection Kit can detect and react specifically to Mycoplasma.
Fig. Specificity of e-Myco™ VALiD-Q Mycoplasma qPCR Detection Kit
A) Mycoplasma hyorhinis(Positive Control), B) Nuclease free water(Negative Control), C) Clostridium perfringens, D) Streptococcus mutans,
E) Lactobacillus plantarum, F) Mobiluncus mulieris, G) Gardnerella vaginalis, H) Chlamydia trachomatis
High sensitivity to meet Mycoplasma negative testing guidelines
Fig. Analytical Sensitivity of e-Myco™ VALiD-Q Mycoplasma qPCR Detection Kit
A) Mycoplasma arginini, B) Mycoplasma fermentans, C) Mycoplasma hyorhinis, D) Mycoplasma orale, E) Mycoplasma pneumoniae, F) Acholeplasma laidlawii.
The e-Myco ™ VALiD-Q Mycoplasma qPCR Detection Kit was able to detect all 6 Mycoplasma types suggested in the guideline. . Through this, the product has the performance in accordance with the Mycoplasma negative test guideline using the nucleic acid amplification method, and it can be confirmed that it is a substitute for the direct method.
Comparative equivalence test with direct method
We tested both the direct method and the sensitivity of this product at the same time. When Mycoplasma was detected by direct incubation using an initial unknown concentration, one or two colonies were observed in samples diluted up to 1/107. When applied to this product, it was confirmed that the target signal was detected up to the concentration of 1/107, which was confirmed by the direct method. This confirms that fraudulent tests using this product can effectively replace the direct method.
Q What is Intron's existing Mycoplasma detection product and e-Myco ™ VALiD-Q Mycoplasma qPCR Detection Kit?
A As a proven product, it is a Mycoplasma detection product that has a sensitivity level that can replace the direct culture method with a detection limit of 10 CFU / ml. In addition, most Mycoplasma can be detected up to 70 kinds and have very high specificity.
Q e-Myco ™ VALiD-Q Mycoplasma What extraction methods can be used to effectively detect Mycoplasma in samples using the QPCR Detection Kit?
A I would recommend the DNA method There is. However, in the case of the cell boiling method, the results may vary depending on the proficiency of the experimenter, so we recommend the Genomic DNA method.
Q Negative control PCR was performed and the target signal was detected. In addition, the experiment was performed using Mycoplasma infection samples, but the internal positive control signal was not measured?
A There are many reasons why a target signal can be detected during analytical control PCR. You may suspect contamination of D.W and test equipment. This product has a high sensitivity to Mycoplasma and the target can be detected even with a small amount of contamination. In addition, the internal control signal is intended to identify errors during testing. If no IPC signal is detected, it is recommended to check PCR condition and template concentration and retest.
|SiZer™-100 DNA Marker Solution||