Scholarlink

<p>Circulating cell-free DNA (cfDNA) has become increasingly important in clinical oncology; however, during routine plasma processing, a substantial proportion of short cfDNA fragments is often lost during extraction. Efficient recovery of short cfDNA fragments from plasma remains a significant technical challenge. We selected an AuNP-based extraction method based on preliminary observations indicating improved retention of short DNA fragments compared with phenol–chloroform, sodium iodide (NaI), and triton–heat–phenol (THP) protocols. The AuNP method leverages the unique surface chemistry of gold nanoparticles, which preferentially bind short DNA fragments via electrostatic interactions and ligand coatings, thereby providing a reliable extraction mechanism. A 171-bp fragment of the BRAF gene was amplified by semi-nested PCR and spiked into plasma samples to simulate cfDNA. DNA recovery was quantified using real-time PCR. The AuNP-based method demonstrated the highest recovery efficiency and sensitivity, with a limit of detection of 10 copies/µL. Conventional methods yielded lower recovery rates and greater variability, particularly at low DNA concentrations. Statistical analysis, based on three independent experiments and performed using one-way ANOVA followed by Tukey’s post hoc test, confirmed significantly improved extraction efficiency for the AuNP-based method compared with all other techniques, with mean recovery increasing from 45% to 85% (95% CI: 80%-90%, p &lt; 0.0001). AuNP-based extraction provides a sensitive and reproducible approach for isolating short cfDNA fragments and may improve downstream molecular analyses of low-abundance DNA targets. Evaluation of this approach using plasma samples from patients with low tumor burden will be necessary to determine its applicability in clinical liquid biopsy workflows.</p> Graphical Abstract <p></p>

错误：搜索内容不能为空，请输入英文关键词
错误：关键词超出字数限制，请精简
高级检索

Gold Nanoparticle–Based Extraction Enhances Recovery and Sensitivity of Circulating Cell-Free DNA for BRAF Mutation Detection

  • 期刊论文

  • 发表日期：

    2026年4月29日

查看全文
  • Noorossadat Seyyedi,
  • Fatemeh Farjadian,
  • Farahnaz Zare,
  • Mozhgan Karimi,
  • Gholamreza Rafiei Dehbidi,
  • Sedigheh Sharifzadeh,
  • Fariba Nikbakht,
  • Abbas Behzad-Behbahani

摘要

Circulating cell-free DNA (cfDNA) has become increasingly important in clinical oncology; however, during routine plasma processing, a substantial proportion of short cfDNA fragments is often lost during extraction. Efficient recovery of short cfDNA fragments from plasma remains a significant technical challenge. We selected an AuNP-based extraction method based on preliminary observations indicating improved retention of short DNA fragments compared with phenol–chloroform, sodium iodide (NaI), and triton–heat–phenol (THP) protocols. The AuNP method leverages the unique surface chemistry of gold nanoparticles, which preferentially bind short DNA fragments via electrostatic interactions and ligand coatings, thereby providing a reliable extraction mechanism. A 171-bp fragment of the BRAF gene was amplified by semi-nested PCR and spiked into plasma samples to simulate cfDNA. DNA recovery was quantified using real-time PCR. The AuNP-based method demonstrated the highest recovery efficiency and sensitivity, with a limit of detection of 10 copies/µL. Conventional methods yielded lower recovery rates and greater variability, particularly at low DNA concentrations. Statistical analysis, based on three independent experiments and performed using one-way ANOVA followed by Tukey’s post hoc test, confirmed significantly improved extraction efficiency for the AuNP-based method compared with all other techniques, with mean recovery increasing from 45% to 85% (95% CI: 80%-90%, p < 0.0001). AuNP-based extraction provides a sensitive and reproducible approach for isolating short cfDNA fragments and may improve downstream molecular analyses of low-abundance DNA targets. Evaluation of this approach using plasma samples from patients with low tumor burden will be necessary to determine its applicability in clinical liquid biopsy workflows.

Graphical Abstract