SN/T 5282-2020 in English
VALIDQuarantine protocol for infection with covert mortality nodavirus
- Issued on:2020-12-30
- Implemented on:2021-07-01
- File Format:PDF
- Delivery:Via email within 1~3 business days
$262.00
Introduction
Analysis of standard technical framework
| Detection method | Sensitivity | Specificity | Time consumption | Applicable scenarios |
|---|---|---|---|---|
| Histopathology | Medium | 85% | 24-48h | Preliminary screening |
| Nested PCR | 102 copies/μL | 98% | 6-8h | Laboratory confirmation |
| Real-time fluorescence RT-PCR | 101 copies/μL | 99% | 3-4h | Rapid quarantine |
| In situ hybridization | Medium | 95% | 24h+ | Pathological localization |
Key Technical Points
1. Sample Processing Specifications
All instruments that come into contact with RNA must be treated with DEPC water. Davidson's AFA fixative is recommended for tissue fixation (see Appendix B for the formula). Muscle and hepatopancreas should be processed separately.
2. Molecular detection optimization
- Primer design: Targeting the 198bp fragment of RNA polymerase gene (real-time fluorescence) and 619bp/165bp fragment (nested PCR)
- Anti-pollution measures: The experimental zoning implements the requirements of SN/T 1193, and it is recommended to use UNG enzyme to prevent carryover contamination
Recommendations for the implementation of the standard
Application case: Quarantine of imported white shrimp from Ecuador
In 2021, Shenzhen Customs adopted this standard to test imported shrimp products with batch number ECU-2103:
① Initial screening found that 3% of the samples had muscle whitening→② Eosinophilic inclusion bodies were found in the tissue pathology→③ Real-time fluorescence RT-PCR detected a Ct value of 28→④ Sequencing confirmed that CMNV was positive→The entire batch of goods was returned.
Technological Evolution
Compared with the 2015 trial method, this standard has the following major upgrades:
• Added real-time fluorescent probe (FAM/BHQ1) to improve detection efficiency
• Optimized the in situ hybridization proteinase K concentration to 10μg/mL
• Clarified the retest rules for the gray area of Ct values (35-40)
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