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Background <p>Conventional PRRSV diagnostic approaches have limitations, including complex laboratory requirements, inability to distinguish active from past infections, and susceptibility to viral mutations.</p> Objectives <p>This study aimed to develop a rapid and specific porcine reproductive and respiratory syndrome virus (PRRSV) detection method using DNA aptamers selected through Cell-SELEX targeting virus-infected cells.</p> Results <p>MARC-145 cells infected with PRRSV strain VR-2332 served as positive targets, whereas uninfected cells were used as negative controls for Cell-SELEX selection using a randomized single-stranded DNA (ssDNA) library. The selected aptamer candidates underwent post-SELEX binding-affinity screening via NanoDrop quantification, secondary structure prediction using M-fold analysis, and specificity validation through an aptamer-based sandwich assay (ABSA) employing swine influenza virus (SIV)-infected cells as heterologous controls. Cell-SELEX yielded 22 distinct aptamer candidates, with three aptamers (PRRSV_apta17, PRRSV_apta18, and PRRSV_apta21) demonstrating superior binding affinity for PRRSV-infected cells. PRRSV_apta17 exhibited optimal thermodynamic stability and highest binding performance. ABSA validation confirmed high specificity for PRRSV-infected cells with minimal cross-reactivity to SIV-infected controls.</p> Conclusion <p>Cell-SELEX successfully generated highly specific DNA aptamers capable of detecting PRRSV-infected cells by recognizing infection-induced extracellular surface modifications. This aptamer-based approach offers promising alternatives for rapid, cost-effective diagnosis of PRRSV with potentially enhanced resistance to viral mutations by targeting conserved host cellular responses, enabling improved point-of-care testing applications.</p>

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Development of an aptamer-based diagnostic method targeting PRRSV-induced cytopathic effects

  • 期刊论文

  • 发表日期：

    2025年12月29日

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  • In-Hwan Oh,
  • Phat-Loc Nguyen,
  • Dae-Young Park,
  • Jin-Pyo Lee,
  • Woo-Ri Shin,
  • Su-Min Lee,
  • Jiho Min,
  • Ji-Young Ahn,
  • Yang-Hoon Kim

摘要

Background

Conventional PRRSV diagnostic approaches have limitations, including complex laboratory requirements, inability to distinguish active from past infections, and susceptibility to viral mutations.

Objectives

This study aimed to develop a rapid and specific porcine reproductive and respiratory syndrome virus (PRRSV) detection method using DNA aptamers selected through Cell-SELEX targeting virus-infected cells.

Results

MARC-145 cells infected with PRRSV strain VR-2332 served as positive targets, whereas uninfected cells were used as negative controls for Cell-SELEX selection using a randomized single-stranded DNA (ssDNA) library. The selected aptamer candidates underwent post-SELEX binding-affinity screening via NanoDrop quantification, secondary structure prediction using M-fold analysis, and specificity validation through an aptamer-based sandwich assay (ABSA) employing swine influenza virus (SIV)-infected cells as heterologous controls. Cell-SELEX yielded 22 distinct aptamer candidates, with three aptamers (PRRSV_apta17, PRRSV_apta18, and PRRSV_apta21) demonstrating superior binding affinity for PRRSV-infected cells. PRRSV_apta17 exhibited optimal thermodynamic stability and highest binding performance. ABSA validation confirmed high specificity for PRRSV-infected cells with minimal cross-reactivity to SIV-infected controls.

Conclusion

Cell-SELEX successfully generated highly specific DNA aptamers capable of detecting PRRSV-infected cells by recognizing infection-induced extracellular surface modifications. This aptamer-based approach offers promising alternatives for rapid, cost-effective diagnosis of PRRSV with potentially enhanced resistance to viral mutations by targeting conserved host cellular responses, enabling improved point-of-care testing applications.