Spinal muscular atrophy (SMA) is a rare autosomal recessive neuromuscular disease. Nusinersen sodium (NS) is the world's first antisense oligonucleotide (ASO) drug for SMA precise targeted therapy. However, the limited half-life of oligonucleotides and their tendency to accumulate in hepatic and renal tissues presented significant challenges for clinical investigation and therapeutic drug monitoring. In this study, we proposed an analytical strategy based on the specific capture of oligonucleotide functionalized fluorescent probes by single stranded binding proteins (SSB) for ultra-sensitive and high-throughput detection of nusinersen sodium in human serum. The magnetic nanoparticles modified with single-strand binding protein (MNPs-SSB) selectively bonded to the red fluorescent quantum dots functionalized with oligonucleotides (RQDs-ssDNA) that were complementary to nusinersen sodium. Upon interaction with nusinersen sodium, RQDs-ssDNA formed a double-stranded complex (RQDs-ssDNA-NS), resulting in enhanced red fluorescence after magnetic separation as it was no longer captured by MNPs-SSB but remained in the supernatant. A quantitative analysis of nusinersen sodium in biological samples was successfully achieved by establishing a relationship between fluorescence intensity and its concentration. The detection signal F/F0 exhibited a linear correlation (R = 0.9871) over a wide range from 0.1 nM to 200 nM, with a limit of detection (LOD) of 0.03 nM, demonstrating the high specificity and rapid analysis time (only 30 min). This method provided a novel approach for sensitive, high-throughput, and specific analysis of nusinersen sodium and similar ASO drugs.

中文翻译：

---

用于检测人血清中努西那森钠的寡核苷酸功能化荧光探针的灵敏分析策略


脊髓性肌萎缩症（SMA）是一种罕见的常染色体隐性遗传神经肌肉疾病。努西那森钠（NS）是全球首个用于SMA精准靶向治疗的反义寡核苷酸（ASO）药物。然而，寡核苷酸有限的半衰期及其在肝和肾组织中积累的趋势给临床研究和治疗药物监测带来了重大挑战。在这项研究中，我们提出了一种基于单链结合蛋白（SSB）特异性捕获寡核苷酸功能化荧光探针的分析策略，用于超灵敏和高通量检测人血清中的nusinersen钠。用单链结合蛋白（MNPs-SSB）修饰的磁性纳米粒子选择性地与用与nusinersen钠互补的寡核苷酸（RQDs-ssDNA）功能化的红色荧光量子点结合。与 nusinersen 钠相互作用后，RQDs-ssDNA 形成双链复合物（RQDs-ssDNA-NS），磁力分离后红色荧光增强，因为它不再被 MNPs-SSB 捕获，而是保留在上清液中。通过建立荧光强度与其浓度之间的关系，成功实现了生物样品中努西那森钠的定量分析。检测信号 F/F0 在 0.1 nM 至 200 nM 的宽范围内呈现线性相关性 (R = 0.9871)，检测限 (LOD) 为 0.03 nM，展示了高特异性和快速分析时间（仅需 30 分钟） ）。该方法为nusinersen钠和类似ASO药物的灵敏、高通量和特异性分析提供了一种新方法。