School of Chemistry, South China Normal University, Guangzhou 51006, China.
Guangzhou Center for Disease Control and Prevention, Guangzhou 510440, China.
Anal Chem. 2021 Oct 26;93(42):14106-14112. doi: 10.1021/acs.analchem.1c01983. Epub 2021 Oct 17.
Dengue fever is caused by mosquito-transmitted dengue virus infection and continues to increase worldwide, threatening public health in tropical and subtropical regions. The primary difficulties in preventing a reduction of the medical burden of dengue fever lies in the lack of effective mosquito control, preventive dengue vaccines, and clinically effective antiviral drugs to treat dengue infections. Rapid and accurate diagnosis is crucial for proper patient care and effective control of epidemics. The present work proposes an alternative strategy for detecting the dengue virus nonstructural protein 1 (NS1) antigen in clinical serum samples by using ultrahigh-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) in combination with the molecularly imprinted polymers. Rather than the whole protein, the NS1 signature peptide is selected as a template for molecular imprinting and quantified as a stoichiometric readout of NS1. Three functional monomers with hydrophobic, positively charged, and negatively charged groups were synthesized by click reactions in terms of the signature peptide. These three functional monomers provide abundant recognition sites for the peptide, allowing the peptide template to be effectively imprinted during polymerization. The imprinting conditions were optimized, and the molecularly imprinted polymers were characterized and used for enriching the signature peptide from digested serum samples by solid-phase extraction and then detected by UHPLC-MS/MS. The proposed method is used to detect the dengue virus NS1 in clinical samples and holds significant promise for early confirmation of dengue virus infection.
登革热是由蚊媒传播的登革病毒感染引起的，在全球范围内持续增加，威胁着热带和亚热带地区的公共卫生。预防登革热医疗负担减轻的主要困难在于缺乏有效的蚊虫控制、预防登革热疫苗和临床有效的抗病毒药物来治疗登革热感染。快速准确的诊断对于患者的适当护理和有效控制疫情至关重要。本工作提出了一种通过超高效液相色谱串联质谱（UHPLC-MS/MS）结合分子印迹聚合物检测临床血清样本中登革病毒非结构蛋白 1（NS1）抗原的替代策略。不是整个蛋白质，而是选择 NS1 特征肽作为分子印迹的模板，并作为 NS1 的化学计量读数进行定量。根据特征肽，通过点击反应合成了具有疏水性、正电性和负电性的三种功能单体。这三种功能单体为肽提供了丰富的识别位点，允许在聚合过程中有效印迹肽模板。优化了印迹条件，并对分子印迹聚合物进行了表征，然后通过固相萃取从消化的血清样本中富集特征肽，再通过 UHPLC-MS/MS 进行检测。该方法用于检测临床样本中的登革病毒 NS1，有望早期确认登革热病毒感染。
