无嘌呤/无嘧啶(AP)位点是最常见的DNA损伤之一,也是碱基切除修复途径中的关键中间体。因此,AP部位在临床诊断中至关重要,治疗和检测。然而,现有的检测方法设计和综合复杂,仪器要求高,限制了其广泛应用。因此,迫切需要一种灵敏和直接的检测方法,而不需要耗时和不均匀的反应。在这里,我们针对长dsDNA和短dsDNA中的AP位点开发了两种兼容的检测方法。对于长和短的dsDNA,末端脱氧核苷酸转移酶(TdT)和3'-末端阻断的亲和力差异成功抑制了背景信号,分别,从而实现高检测性和特异性。检测限为13pM在20μL,这意味着AP位点数量的LOD为0.26fmol,AP位点丰度的LOD为0.05%。该方法已成功应用于各种生物样品中AP位点的快速检测。因此,具有广阔的临床应用前景,迎合了一点护理的需要。
The apurinic/apyrimidinic (AP) site is one of the most common DNA lesions and a critical intermediate during the base excision repair pathway. Therefore, AP sites are essential in clinical diagnosis, treatment and detection. However, the existing detection methods are complicated in design and synthesis and have high instrument requirements, limiting their wide application. Therefore, there is an urgent need for a sensitive and straightforward detection method without time-consuming and heterogeneous reactions. Herein, we developed two compatible detection methods for AP sites in long and short dsDNA. For long and short dsDNA, the background signal was successfully suppressed by the affinity difference of Terminal deoxynucleotidyl transferase (TdT) and 3\' -end blocking, respectively, thus achieving high detectability and specificity. The detection limit was 13 pM in 20 μL, meaning that the LOD was 0.26 fmol for AP site amount and 0.05% for AP site abundance. The method has been successfully applied to detect AP sites in various biological samples quickly. Therefore, it has broad clinical application prospects, catering for the need for a point of care.