SN/T 5484-2022 in English

Analysis of standard technical framework

This standard establishes a composite detection system that combines virus isolation and culture in Striped salmon cell line (E-11) with molecular detection, which includes the following core modules:

Key Technical Points

1. Standardized Process for Virus Isolation

The E-11 cell line was used for virus culture, and the standard requirements were:

• Sample processing: Differential dissection of fish of different body lengths (viscera from whole fry, liver/brain/kidney/spleen from adult fish)
• Culture conditions: 25±2℃ closed culture, CPE determination requires simultaneous setting of positive and blank controls
• Passage requirements: Blind culture is required for primary culture without CPE for 7 days, and freeze-thaw treatment improves the detection rate

2. Innovative design of molecular detection

The standard innovatively adopts a double insurance detection strategy:

Implementation and application suggestions

1. Laboratory configuration requirements

• Biological safety cabinet must reach Class II A2 type
• Fluorescence PCR instrument must have FAM channel
• Cell culture requires a 25℃ constant temperature CO₂ incubator

2. Key points for result interpretation

Suspicious case judgment must meet the following requirements:
Ct value ≤35 and obvious logarithmic phase of amplification curve, or 250bp band appears in nested PCR

3. Cross-border quarantine application

It is recommended to implement the following for live tilapia from epidemic areas:
Triple testing (clinical symptom observation + virus isolation + molecular testing)