Synthesis, Luminescence Properties Of N-(aminobutyl)-N-(ethylisoluminol)Functionalized Nanomaterials And Their Applications In Bioassays

In this dissertation, the state of arts in the field of chemiluminescence (CL), chemiluminescent functionalized nanomaterials (CF-NMs) and their applications in bioassays were reviewed. Recently, much attention has been paid to CF-NMs and a series of CF-NMs have been synthesized. The analytical method based on CL-NMs as analytical probes and analytical interface is developed rapidly, showing promising application potential in clinical diagnosis, food safety, environmental monitoring etc. However, the CL efficiency of CF-NMs is limited and it remains great challenge for the detection of trace substance in the biological systems. The aim of this dissertation is to explore the synthesis of CF-NMs with high CL yield and their applications in label-free bioassays. N-(aminobytyl)-N-(ethylisoluminol)(ABEI) functionalized gold nanopopcorns and ABEI functionalized gold nanodots onto chitosan-grafted multi-walled carbon nanotubes (ABEI-AuNDs-cs-MWCNTs) were successfully synthesized. These nanocomposites exhibited excellent CL and electrochemiluminescence (ECL) properties. Based on the as-prepared CF-NMs, novel label-free ECL biosensors for highly sensitive and selective detection of mercury and N-terminal of the prohormone brain natriuretic peptide (NT-proBNP) were developed based on the target-induced directed ECL signal change. Besides, the fluorescence property of ABEI functionalized gold nanoparticles (ABEI-AuNPs) and applications of ABEI-AuNPs in fluorescence resonance energy transfer were studied. The main results are as follows:1. A facile method for the synthesis of popcorn-shaped gold nanomaterials (GNMs) with high CL yield and good monodispersity by reducing LA and HAuCl4with ABEI in ethyl alcohol solution at room temperature through a seed growth method was reproted. The morphologies of GNMs were characterized by transmission electron microscopy, scanning electron microscopy and energy dispersive spectroscopy. The results showed that various morphologies of GNMs from gold nanopopcorns (AuNPCs), quasi-spherical to irregular GNMs could be obtained. The surface composition of AuNPCs was studied and the results strongly supported that ABEI, the oxidation product of ABEI and reduction product of LA were attached to a gold core by covalent bonds. A possible mechanism of shape evolution is proposed. The special morphology of AuNPCs maintains the large specific surface area and provides excellent catalytic property when involved in the CL reaction. Thus the CL functionalized AuNPCs exhibited good CL property (two orders of magnitude higher than that of ABEI functionalized gold nanoparticles in our previous work). Furthermore, DNA and protein labeled with AuNPCs showed excellent CL activity, which implies their application potential as bio-probes for immunoassay and DNA assay.2. A one-step strategy for high-density assembly of ABEI functionalized gold nanodots onto the sidewalls of chitosan-grafted multi-walled carbon nanotubes (ABEI-AuNDs-cs-MWCNTs) via the reduction of HAuCl4with ABEI in the presence of cs-MWCNTs. During the synthesis process, a great number of ABEI molecules as stabilizers were directly coated on the surface of ABEI-AuNDs-cs-MWCNTs. The morphology and surface composition of ABEI-AuNDs-cs-MWCNTs were studied by transmission electron microscopy, energy dispersive spectroscopy, X-ray photoelectron spectroscopy and Raman spectra. The results showed that ABEI-AuNDs uniformly distributed on the surface of composites. Furthermore, the CL and ECL properties of ABEI-AuNDs-cs-MWCNTs were studied and ABEI-AuNDs-cs-MWCNTs exhibited excellent CL and ECL properties. Based on the excellent ECL activity of ABEI-AuNDs-cs-MWCNTs and the good electro-transfer property of MWCNTs and film-forming property of chitosan, a novel label-free ECL biosensor was developed and has an extremely sensitive response to mercury ions in a liner range of1.0×10-6mol/～1.0×10-8mol/L, and the detection limit was3.2×10-9mol/L.3. A novel lable-free immunoassay for NT-proBNP was developed based on the ABEI-AuNDs-cs-MWCNTs as ECL sensing platform. ITO-coated glass slides were used as an assembly electrode owing to their stability, easy miniaturization and compatibility with microfabrication. After a centrifugation-ultrasonic dispersion process, ABEI-AuNDs-cs-MWCNTs were dripped onto the surface of ITO glass and ABEI-AuNDs-cs-MWCNTs functionalized ITO electrode was obtained. Glutaraldehyde molecules were used as bridge molecules for anti-NT-proBNP coated on the surface of ITO electrode. After blocking with bovine serum albumin, the electrode could be used as a label-free ECL immunosensor for the detection of NT-proBNP. ECL behavior of the modified electrode was studied by a homemade ECL system. It demonstrated that the ABEI-AuNDs-cs-MWCNTs constructed nano-interface showed high ECL intensity. The as-prepared label-free ECL immunosensor has an extremely sensitive response to NT-proBNP in a liner range of1.0×10-10g/mL-1.0×10-14g/mL with the detection limit of3.86×10-15g/mL. It also shows good precision, acceptable stability and reproducibility. This work provides a promising technique for protein detection, which may open up a new avenue for the ultrasensitive label-free immunoassays.4. The FL properties of as-prepared ABEI-AuNPs were studied in this work. It was observed that as-prepared ABEI-AuNPs with a concentration of1.23×10-9mol/L displayed comparable FL emission to that of the7.8×10-7mol/L ABEI solution. Although86%of the FL emission of ABEI molecules on the surface of ABEI-AuNPs was inhibited by gold cores through intra-and inter-particle quenching effects, the FL intensity of a gold nanoparticle was still-634times brighter than that of a single ABEI molecule. The as-prepared ABEI-AuNPs also exhibited much better photostability than luminol-AuNPs reported by our previous work.96%FL intensity of as-prepared ABEI-AuNPs solution remained after5min continuous excitation and only80%FL intensity of luminol-AuNPs would be kept in our previous work. Furthermore, almost70%FL intensity of ABEI-AuNPs still remained after45min. Moreover, ABEI-AuNPs and acriflavine were successfully used as energy donor and energy acceptor, respectively, to construct a novel FRET system. And an efficient energy transfer from ABEI-AuNPs to acriflavine in the solution was observed. This work demonstrates for the first time that ABEI functionalized gold nanomaterials could be used as energy donor. Taking advantage of excellent labeling property of gold nanomaterials, the FRET property between ABEI-AuNPs and acriflavine may find future applications in bioassays with FL detection.