Key Laboratory of Agricultural Information Acquisition Technology, Ministry of Agriculture, China Agricultural University, Beijing, China.
Key Laboratory on Modern Precision Agriculture System Integration Research, Ministry of Education, China Agricultural University, Beijing, China.
Theranostics. 2018 Nov 29;8(22):6263-6273. doi: 10.7150/thno.29025. eCollection 2018.
In this study, an enzyme-free biosensor was developed for sensitive and specific detection of Salmonella typhimurium (S. typhimurium) using curcumin (CUR) as signal reporter and 1,2,4,5-tetrazine (Tz)-trans-cyclooctene (TCO) click chemistry for signal amplification.
Nanoparticles composed of CUR and bovine serum albumin (BSA) were formulated and reacted with Tz and TCO to form Tz-TCO-CUR conjugates through Tz-TCO click chemistry. Then, the Tz-TCO-CUR conjugates were functionalized with polyclonal antibodies (pAbs) against S. typhimurium to form CUR-TCO-Tz-pAb conjugates. Magnetic nanoparticles (MNPs) conjugated with monoclonal antibodies (mAbs) against S. typhimurium through streptavidin-biotin binding were used to specifically and efficiently separate S. typhimurium from the background by magnetic separation. CUR-TCO-Tz-pAb conjugates were reacted with the magnetic bacteria to form CUR-Tz-TCO bacteria. Finally, CUR was released quickly from the CUR-Tz-TCO bacteria in the presence of NaOH, and the color change was measured at the characteristic wavelength of 468 nm for bacteria quantification.
A linear relationship between absorbance at 468 nm and concentration of S. typhimurium from 102 to 106 CFU/mL was found. The lower detection limit was calculated to be as low as 50 CFU/mL and the mean recovery was 107.47% for S. typhimurium in spiked chicken samples.
This biosensor has the potential for practical applications in the detection of foodborne pathogens.
本研究开发了一种无酶生物传感器，用于灵敏和特异性检测鼠伤寒沙门氏菌（Salmonella typhimurium，S. typhimurium），使用姜黄素（CUR）作为信号报告物，并使用 1,2,4,5-四嗪（Tz）-反式环辛烯（TCO）点击化学进行信号放大。
制备由 CUR 和牛血清白蛋白（BSA）组成的纳米颗粒，并与 Tz 和 TCO 反应，通过 Tz-TCO 点击化学形成 Tz-TCO-CUR 缀合物。然后，Tz-TCO-CUR 缀合物与针对 S. typhimurium 的多克隆抗体（pAbs）反应，形成 CUR-TCO-Tz-pAb 缀合物。通过链霉亲和素-生物素结合将针对 S. typhimurium 的单克隆抗体（mAbs）偶联到磁性纳米颗粒（MNPs）上，通过磁分离特异性和有效地从背景中分离 S. typhimurium。CUR-TCO-Tz-pAb 缀合物与磁性细菌反应，形成 CUR-Tz-TCO 细菌。最后，在 NaOH 的存在下，CUR 从 CUR-Tz-TCO 细菌中快速释放，并且在细菌定量的特征波长 468nm 处测量颜色变化。
在 102 到 106 CFU/mL 的范围内，在 468nm 处的吸光度与 S. typhimurium 浓度之间存在线性关系。检测下限计算低至 50 CFU/mL，在添加鸡肉样品中的 S. typhimurium 时的平均回收率为 107.47%。
该生物传感器具有在检测食源性病原体中的实际应用潜力。
