African Swine Fever Virus (Internal Control) Real Time PCR Detection Kit

REF: GZS001| SKU: 317339| BRAND: GZ MED

Scientific Overview

The African Swine Fever Virus (Internal Control) Real Time PCR Detection Kit is an advanced molecular diagnostic support tool designed for the qualitative detection of African Swine Fever Virus (ASFV) nucleic acid. Utilizing Real-Time PCR technology coupled with the $5^{\prime}\rightarrow3^{\prime}$ exonuclease activity of Taq DNA polymerase, the kit offers diagnostic detection support with high specificity and sensitivity through hydrolysis fluorogenic probes. The system incorporates specific primers and probes targeting conserved regions of the swine genome as an Internal Control, enabling simultaneous monitoring of sampling, DNA extraction, and amplification stages to prevent false negatives. Target ASFV amplification is detected in the FAM channel, while the Internal Control is monitored via the HEX channel. This kit serves as an efficient diagnostic support for rapid detection, clinical evaluation, and epidemiological screening both in field conditions and laboratory environments for professional veterinary use.

Execution Protocol

• Sample Preparation:
  • Whole Blood / Serum: Mix 200 µL of sample with preservation buffer, then extract 200 µL for processing.
  • Swabs (oral, nasal, anal, environmental): Insert the collected swab into preservation buffer, vortex thoroughly to dissociate pathogens, and collect 200 µL for extraction.
  • Tissue Samples (spleen, lymph nodes, etc.): Homogenize 50 mg of tissue in 0.5 mL PBS or preservation buffer (10% homogenate). Centrifuge at 5000 rpm for 5 minutes and retrieve 200 µL of supernatant.
• Automated Nucleic Acid Extraction Phase: Take out the pre-filled single-use magnetic bead cartridge and peel off the protective sealing film. Transfer 200 µL of prepared sample into the loading well labeled “LB”. Place the conductive pipette tip into the first position of the extraction cartridge, situated to the left of the “LB” well.
• Cycle Automation: Insert the cartridge into the extraction compartment of the VET PCR system, confirming activation of the red LED indicator on the corresponding channel. Initiate the protocol: the automated purification cycle completes in approximately 8 minutes.
• PCR Reaction Setup: Carefully collect 20 µL of purified nucleic acid (eluate) from the final “EB” well and transfer it directly into the tube containing the GZ MED lyophilized master mix (compact white cake).
• Reconstitution and Loading: Fully reconstitute the lyophilized master mix by gently tapping the bottom of the tube with fingers. Ensure through gentle flicking or brief centrifugation (10 seconds) that the total volume settles at the bottom and no air bubbles remain, preventing interference with background fluorometric signals.
• Automated Result Interpretation: Place the reaction tubes into the Real-Time PCR instrument and execute the specified thermal profile. The software automatically analyzes amplification curves:
  • ASFV Positive: FAM channel with Ct < 36 and a typical sigmoidal S-curve.
  • ASFV Negative: FAM channel with Ct = 0 and HEX channel with Ct < 36 (sample validity confirmation). *Note: For environmental swabs, lack of amplification in the HEX channel is considered normal; evaluation relies exclusively on the FAM channel.
  • Borderline Result: FAM channel with 36 ≤ Ct < 40. Retesting is required.
  • Invalid Test: FAM channel with Ct = 0 and HEX channel with Ct = 0. Indicates PCR failure; the session must be repeated.

Performance Specifications

Technical opinion: Implementing a Real-Time PCR diagnostic support system with integrated endogenous internal control represents a pivotal step for biosecurity management and epidemiological surveillance in swine farming. The native workflow optimization on the Gangzhu Medical PCR VET system eliminates thermal degradation risks through single-dose lyophilized master mixes and significantly reduces operational time by isolating nucleic acid in just 8 minutes. As a diagnostic support tool designated for professional use only, any negative result does not categorically exclude infection and must be interpreted and contextualized by the Veterinary Surgeon alongside overall clinical examination, case history, and applicable health surveillance regulations.

BIOLAB Diagnostici Technical Team – Veterinary Division

Biolab is the exclusive distributor in Italy for GZ MED