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Real-time PCR Detection Kit for Peste des Petits Ruminants Virus (PPRV)
【Product Name】Real-time PCR Detection Kit for Peste des Petitis Ruminants Virus(PPRV)
【Package】50 Tests
【Indication】The Real-time PCR Detection Kit for Peste des Petitis Ruminants Virus(PPRV) is applicable to detect the Peste des Petitis Ruminants Virus(PPRV) RNA in animal blood, tissue, ulcer fluid and cell culture samples. The test results are for research purposes only and not for clinical diagnosis.
【Main components and content】
Name | Specification | Quantity |
PPRV Reaction Solution | 1000µl/Tube | 1 |
PPRV Positive Control | 250µl/Tube | 1 |
Negative Control | 250µl/Tube | 1 |
【Storage and Shelf Life】
Stored at -20±5℃,Repeated freezing and thawing ≤ 3 times, the shelf life is 12 months.
【Test Method】
1. Nucleic Acid Extraction
A commercialized RNA/DNA extraction kit can be carried out for nucleic acid extraction, please follow the instruction of the kit.
2. PCR amplification
2.1 Calculate the number of test samples, take n+2 PCR reaction tubes, and add 20µl to of reaction solution to each tube.
2.2 Add 5µl nucleic acid of negative control, positive control and samples into the above PRC reaction tubes respectively, centrifuge at 8000rpm for several seconds, and put them into the fluorescent quantitative PCR amplifier.
2.3 The reaction conditions shall be set as:
Parameters of amplification | |||
System | Total volume is 25µl | ||
Signal collection | PRRV fluorescence signal | FAM channel collects fluorescence signal | |
Reaction Conditions of PCR | Phase | Conditions | Cycles |
UNG Process | 55℃ for 15 minutes | 1 | |
Pre-denaturation | 95℃ for 30 seconds | 1 | |
PCR | 95℃ for 10 seconds | 40 | |
60℃ for 30 seconds Set to collect the fluorescent signal at the end of this phase | |||
【Interpretation of the results】
1. The validity judgement:
1.1 Positive Control: The Ct value of FAM channel ≤ 32, and the amplification curve has a significant exponential growth phase.
1.2 Negative Control: FAM channel has not amplification curve, or the amplification curve is straight line or slight oblique line.
2. The test result judgement:
If the FAM detection channel of the test sample has no amplification curve, it can be determined as PPRV negative.
If the FAM detection channel has an exponential growing amplification curve and the Ct value is ≤ 36, it can be determined as PPRV positive.
36<Ct Value<40 samples should be considered as doubtful results, and the analysis should be repeated for confirmation.
【Precautions】
1. The laboratory management shall strictly follow the management specification of PCR gene amplification laboratory. The laboratory personnel must be trained professionally. The experiment process shall be conducted strictly in different areas (Reagent preparation area, specimen preparation area, amplification and product analysis area). All consumables shall be disposable after sterilization. Special apparatus, equipment and supplies at each stage of the experiment operation shall not be cross-used.
2. Please prepare the biological safety cabinet for reagent and specimen preparation stage. The Lab coat, disposable gloves and pipette shall be carried out during the experiment.
3. Repeated freezing&thawing of reagents shall be avoided as far as possible. Before use, the reagents shall be completely thawed and centrifuged at 8000rpm for several seconds.
4. Please put the pipette used in the specimen preparation area into the container containing disinfectant and discard with the waste after sterilization.
5. After the experiment, the worktable and pipette shall be treated with 10% hypochlorite or 75% alcohol or ultraviolet lamp.
【Manufacture】
Name: Shandong Xinda Gene Technology Co., Ltd
A subsidiary of the Shandong Sinder Technology Co., Ltd
Address: Building B2, Bandaohuigu Industrial Park, Shungeng Road, Zhucheng City, Shandong Province
Post Code: 262233
Phone: +86 - 0532 5882 0810
Real-time PCR Detection Kit for Peste des Petits Ruminants Virus (PPRV)
【Product Name】Real-time PCR Detection Kit for Peste des Petitis Ruminants Virus(PPRV)
【Package】50 Tests
【Indication】The Real-time PCR Detection Kit for Peste des Petitis Ruminants Virus(PPRV) is applicable to detect the Peste des Petitis Ruminants Virus(PPRV) RNA in animal blood, tissue, ulcer fluid and cell culture samples. The test results are for research purposes only and not for clinical diagnosis.
【Main components and content】
Name | Specification | Quantity |
PPRV Reaction Solution | 1000µl/Tube | 1 |
PPRV Positive Control | 250µl/Tube | 1 |
Negative Control | 250µl/Tube | 1 |
【Storage and Shelf Life】
Stored at -20±5℃,Repeated freezing and thawing ≤ 3 times, the shelf life is 12 months.
【Test Method】
1. Nucleic Acid Extraction
A commercialized RNA/DNA extraction kit can be carried out for nucleic acid extraction, please follow the instruction of the kit.
2. PCR amplification
2.1 Calculate the number of test samples, take n+2 PCR reaction tubes, and add 20µl to of reaction solution to each tube.
2.2 Add 5µl nucleic acid of negative control, positive control and samples into the above PRC reaction tubes respectively, centrifuge at 8000rpm for several seconds, and put them into the fluorescent quantitative PCR amplifier.
2.3 The reaction conditions shall be set as:
Parameters of amplification | |||
System | Total volume is 25µl | ||
Signal collection | PRRV fluorescence signal | FAM channel collects fluorescence signal | |
Reaction Conditions of PCR | Phase | Conditions | Cycles |
UNG Process | 55℃ for 15 minutes | 1 | |
Pre-denaturation | 95℃ for 30 seconds | 1 | |
PCR | 95℃ for 10 seconds | 40 | |
60℃ for 30 seconds Set to collect the fluorescent signal at the end of this phase | |||
【Interpretation of the results】
1. The validity judgement:
1.1 Positive Control: The Ct value of FAM channel ≤ 32, and the amplification curve has a significant exponential growth phase.
1.2 Negative Control: FAM channel has not amplification curve, or the amplification curve is straight line or slight oblique line.
2. The test result judgement:
If the FAM detection channel of the test sample has no amplification curve, it can be determined as PPRV negative.
If the FAM detection channel has an exponential growing amplification curve and the Ct value is ≤ 36, it can be determined as PPRV positive.
36<Ct Value<40 samples should be considered as doubtful results, and the analysis should be repeated for confirmation.
【Precautions】
1. The laboratory management shall strictly follow the management specification of PCR gene amplification laboratory. The laboratory personnel must be trained professionally. The experiment process shall be conducted strictly in different areas (Reagent preparation area, specimen preparation area, amplification and product analysis area). All consumables shall be disposable after sterilization. Special apparatus, equipment and supplies at each stage of the experiment operation shall not be cross-used.
2. Please prepare the biological safety cabinet for reagent and specimen preparation stage. The Lab coat, disposable gloves and pipette shall be carried out during the experiment.
3. Repeated freezing&thawing of reagents shall be avoided as far as possible. Before use, the reagents shall be completely thawed and centrifuged at 8000rpm for several seconds.
4. Please put the pipette used in the specimen preparation area into the container containing disinfectant and discard with the waste after sterilization.
5. After the experiment, the worktable and pipette shall be treated with 10% hypochlorite or 75% alcohol or ultraviolet lamp.
【Manufacture】
Name: Shandong Xinda Gene Technology Co., Ltd
A subsidiary of the Shandong Sinder Technology Co., Ltd
Address: Building B2, Bandaohuigu Industrial Park, Shungeng Road, Zhucheng City, Shandong Province
Post Code: 262233
Phone: +86 - 0532 5882 0810
