Abstract：
Exosomal glycoproteins play a significant role in many physiological and pathological processes. However, the detection of exosome surface glycans is currently challenged by the complexity of biological samples or the sensitivity of the methods. Herein, we prepared a novel fluorescent probe of biotin-functionalized nanocrystals (denoted as CdTe@cys-biotin) and applied it for the first time for the detection of the expression of exosomal surface glycans using a fluorescence amplification strategy. First, the dual affinity of TiO2 and CD63 aptamers of Fe3O4@TiO2-CD63 was utilized to rapidly and efficiently capture exosomes within 25 min. In this design, interference from other vesicles and soluble impurities can be avoided due to the dual recognition strategy. The chemical oxidation of NaIO4 oxidized the hydroxyl sites of exosomal surface glycans to aldehydes, which were then labeled with aniline-catalyzed biotin hydrazide. Using the high affinity between streptavidin and biotin, streptavidin-FITC and probes were successively anchored to the glycans on the exosomes. The fluorescent probe achieved the dual function of specific recognition and fluorescent labeling by modifying biotin on the surface of nanocrystals. This method showed excellent specificity and sensitivity for exosomes at concentrations ranging from 3.30 × 102 to 3.30 × 106 particles/mL, with a detection limit of 121.48 particles/mL. The fluorescent probe not only quantified exosomal surface glycans but also distinguished with high accuracy between serum exosomes from normal individuals and patients with kidney disease. In general, this method provides a powerful platform for sensitive detection of exosomes in cancer diagnosis.
摘要：
外泌体糖蛋白在许多生理和病理过程中起着重要作用。然而,外来体表面聚糖的检测目前受到生物样品的复杂性或方法的敏感性的挑战。在这里，我们制备了一种新型的生物素功能化纳米晶体荧光探针（称为CdTe@cys-生物素），并首次使用荧光扩增策略将其用于检测外泌体表面聚糖的表达。首先,利用Fe3O4@TiO2-CD63的TiO2和CD63适体的双重亲和力在25分钟内快速有效地捕获外泌体。在这个设计中，由于双重识别策略，可以避免来自其他囊泡和可溶性杂质的干扰。NaIO4的化学氧化将外泌体表面聚糖的羟基位点氧化为醛，然后用苯胺催化的生物素酰肼标记。利用链霉亲和素与生物素的高亲和力,链霉亲和素-FITC和探针相继锚定到外泌体上的聚糖。该荧光探针通过在纳米晶表面修饰生物素,实现了特异性识别和荧光标记的双重功能。该方法对浓度范围为3.30×102至3.30×106颗粒/mL的外来体显示出优异的特异性和灵敏度，检测限为121.48颗粒/mL。荧光探针不仅定量了外泌体表面聚糖，而且还高精度地区分了正常人和肾病患者的血清外泌体。总的来说,该方法为癌症诊断中外泌体的灵敏检测提供了强大的平台。
