Multifunctional Nanospheres For Rapid Point-of-Care Detection Of Infectious Disease

Infectious diseases continue to be a major threat to human health and property security.Early,rapid and robust detection of highly contagious pathogens is the key to early diagnosis,treatment,and prognosis of diseases.It is also significance to increase the probability of survival and reduce infection rates.Lateral flow immunoassay is currently the most promising format for point-of-care diagnosis,since its advantages included portability,speed,cost-effectiveness,and ease-of-use.With the development and deep explorations of nanoscience and nanotechnology,various nanomaterials have been applied to the lateral flow immunoassays and showed great superiority and potential.Among them,gold nanoparticles(Au NPs),quantum dots(QDs)and magnetic materials have showed excellent performance in the field due to their incomparable properties.In the past decade,our group has always been committed to the construction,research,and application of QDs and magnetic materials.Based on these materials,our group has developed two methods to fabricate fluorescent/magnetic nanospheres.Take advantage of the fluorescent/magnetic nanospheres,our group has realized sensitive and reliable detection of proteins,virus,bacteria and tumor cells.In this thesis,we have developed multifumctional nanospheres based lateral flow immunoassays,and applied them to the detection of protein marker,virus,and bacterium,which have been summarized as follows:(1)A quantitative lateral flow immunoassay with high sensitivity for C-reaction protein(CRP)was established by utilizing fluorescent nanospheres(FNs),which was fabricated by embedding QDs into polymer nano spheres.The FNs exhibited superstrong fluorescence,380-fold higher than that of one QD.Due to the excellent fluorescence properties of FNs,a detection limit of 27.8 pM CRP was achieved within 20 min.Moreover,other cancer biomarkers showed negative results by this method,validating the excellent specificity of the assay.Then its successfully application to the peripheral blood plasma samples from lung cancer and breast cancer patients,and healthy people,indicating its clinical application values.Moreover,The FNs based lateral flow immunoassay holds a good prospect of CRP detection in clinical application and can be expanded to other protein biomaker detection.(2)To develop an EBOV glycoprotein-based POC assay,we designed an enhanced dual-signal readout lateral flow immunoassay with a novel reporter(RNs@Au).It was fabricated by adhering Au NPs onto the surface of the red fluorescent nanospheres(RNs).Each RNs@Au contains approximately hundreds of quantum dots and dozens of Au NPs,which enhance both the fluorescence and the colorimetric signals compared to a QD or Au NP.Then,the biotin-streptavidin system was introduced to fuether amplify the dual signal.With the enhanced assay,as low as 2 ng/mL EBOV glycoprotein can be visualized with the naked eye within 20 min,and the detection limit is 0.18 ng/mL.Additionally,the assay was successfully applied for quantitative detection of the EBOV glycoprotein,as well as inactivated whole virion of complex samples in the field only with a smartphone.The enhanced RNs@Au-based lateral flow immunoassay is very convenient,reliable,sensitive,and time-saving,holding great promising in the EBOV POC detection in the field and clinics,especially in resource-poor areas.The merits of the novel reporter,dual-signal readout,ability for quantitation,high sensitivity,and anti-interference may substantially broaden the applications of lateral flow assays for early detection of disease biomarkers and virus.(3)A novel reporter of lateral flow immunoassay was fabricated by assembling red QDs and magnetic nanoparticles onto polymer nanospheres.The obtained fluorescent-magnetic nanospheres(RMNs)can provide three signals(colorimetric signal,fluorescence signal and magnetic signal),and can meet the requirements of different medical conditions.Moreover,the introduction of magnetic separation realized the separation and enrichment of Salmonella typhimurium as well as increased the anti-interfernence ability of the assay.It also increased the sensitivety by one of magnitude.With the assay,as low as 1.88×104 CFU/mL Salmonella typhimurium can be visualized with the naked eye and 3.75×103 CFU/mL Salmonella typhimurium can be detected with a magnetic strip reader.In addition,the assay realized two formats of quantitative detection by measuring the fluorescence and magnetic signal,and the range of lineary is 1.88×104-1.88×107 CFU/mL.Furthernore,its successful application to the spiked samples(milk,fetal bovine serum and blood)had validated its potentiality in real,complex sample detection.The multi-signal readout of the fluorescent-magnetic nanospheres holds great promising prospect of application,especially in fields such as clinical diagnosis,food safety,environmental monitoring and forth.