The dried blood spot (DBS) samples are a useful resource for viral DNA isolation and important in increasing access to HBV diagnosis. However, the choice of the DNA extraction method is crucial for reliable results. We compared the reliability of four DNA extraction methods using DBS samples for the qualitative and quantitative detection of HBV. A panel of serially diluted HBV DNA in whole blood was spotted onto filter paper (Whatman 903 paper and Whatman FTA cards). Four methods were used to extract DNA: QIAamp^® DNA Blood Mini Kit (Qiagen); High Pure Viral Nucleic Acid Kit (Roche); Invisorb Spin Blood Midi Kit (Invitek), and DBS Genomic DNA Isolation Kit (Norgen Biotek). Two qualitative PCRs for the core and surface gene regions of HBV were used, and in‐house real‐time PCR was also evaluated. It was possible to detect HBV DNA using all extraction and PCR protocols. The lowest limit of detection was found using Whatman 903 paper, Roche extraction, and qualitative PCR (20 copies of HBV DNA per ml) for the surface/polymerase HBV region. In the case of in‐house real‐time PCR, the lowest limit of detection was found using both Roche and Qiagen assays (estimated 2 × 10³ copies per ml). These results suggest the importance of both the extraction method and PCR protocol in detecting HBV DNA in DBS. This study provides insights into the utility of DBS samples in HBV molecular diagnosis and their feasibility in low resource areas where cold storage and transportation may be difficult.

中文翻译：

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四种提取方法检测干血斑样乙肝病毒DNA的比较

干血斑 (DBS) 样本是病毒 DNA 分离的有用资源，对于增加获得 HBV 诊断的机会很重要。然而，DNA 提取方法的选择对于获得可靠的结果至关重要。我们比较了四种 DNA 提取方法使用 DBS 样本进行 HBV 定性和定量检测的可靠性。在滤纸（Whatman 903 纸和 Whatman FTA 卡）上点涂了一组在全血中连续稀释的 HBV DNA。使用四种方法提取 DNA： QIAamp ^®DNA 血液迷你试剂盒（Qiagen）；高纯病毒核酸试剂盒（罗氏）；Invisorb Spin Blood Midi Kit (Invitek) 和 DBS Genomic DNA Isolation Kit (Norgen Biotek)。使用了 HBV 核心和表面基因区域的两种定性 PCR，并评估了内部实时 PCR。可以使用所有提取和 PCR 协​​议检测 HBV DNA。使用 Whatman 903 论文、Roche 提取和定性 PCR（每毫升 20 个 HBV DNA 拷贝）发现了表面/聚合酶 HBV 区域的最低检测限。在内部实时 PCR 的情况下，使用 Roche 和 Qiagen 检测（估计 2 × 10 ³每毫升份数）。这些结果表明提取方法和 PCR 方案在 DBS 中检测 HBV DNA 中的重要性。这项研究提供了对 DBS 样本在 HBV 分子诊断中的效用及其在冷藏和运输可能困难的资源匮乏地区的可行性的见解。