Detecting Intact Virus Using Exogenous Oligonucleotide Labels.

Area of Science:

• Virology
• Molecular Biology
• Diagnostic Assays

Background:

• The COVID-19 pandemic highlighted the need for scalable viral testing solutions.
• Traditional RT-qPCR sensitivity decreases significantly with increased sample pooling.
• Existing methods struggle with sample dilution challenges in pooled testing.

Purpose of the Study:

• To introduce a novel method, DIVER, for quantifying intact virus.
• To assess DIVER's robustness against sample dilution and pooling.
• To evaluate DIVER's potential for efficient viral detection in pooled samples.

Main Methods:

• Detection of intact virus by exogenous-nucleotide reaction (DIVER) involves tagging virus particles with exogenous oligonucleotides.
• Tagged particles are captured using virus-specific agents (e.g., ACE2) on functionalized beads.
• Quantification is achieved by measuring oligonucleotide tags via qPCR.

Main Results:

• DIVER demonstrated detection of 1 × 10^5 liposomes and 100 PFU of lentivirus.
• The method successfully identified positive samples within pooled experiments.
• DIVER shows resilience to sample dilution, a key advantage for pooling.

Conclusions:

• DIVER provides a sensitive and dilution-robust method for intact virus quantification.
• This assay is well-suited for efficient sample pooling strategies.
• DIVER shows promise for clinical validation in viral testing scenarios.