Graphene And Two-dimensional Graphene Analogues For Luminol Chemiluminescence Bioassays

In recent years,chemiluminescence?CL?has been widely applied in biomedicine due to its high sensitivity,wide liner dynamic range,fast analysis and simple equipment.Among many well-established CL systems,luminol is the most extensively studied CL system on account of the fairly high quantum yield and the good water solubility.With the appearance of nanomaterials,especially two-dimensional?2D?graphene,CL has been developed rapidly.However,There were very few reports concerning long-persistent CL functionalized hybrids and CL biosensors based on 2D graphene analogues.In this paper,2D graphene and graphene analogues were used as analytical platform to synthesize a long-persistent chemiluminescent hybrid and develop a series of novel luminol CL bioassays based on 2D graphene and graphene analogues for biomolecules detection.?1?A novel and facile approach was designed to prepare luminol and gold nanoparticles?AuNPs?co-functionalized reduced graphene oxide?rGO?hybrids?rGO/AuNPs/luminol?with long duration CL emission,and further developed for cholesterol detection.Luminol and positively charged AuNPs were absorbed on rGO via?-?stacking and electrostatic interaction,respectively.The as-prepared hybrids could initiate long-persistent CL without decay for one hour when reacted with oxidant H₂O₂under alkaline condition.The mechanism of long-persistent CL may attribute to the synergetic catalysis of rGO and AuNPs.It not only promoted a high yield of radical formation,but also improved the stability of radical.Owing to large?–network plane of rGO,^·OH and O₂^·-,producing by the decomposion of H₂O₂,would further react to form ¹O₂ at the surface of rGO.In addition,^·OH could add to double bonds at the rGO plane to generate abundant oxidizing?–conjugated carbon radicals??–C=C^·?,which could directly inducing long-persistent CL.As cholesterol in the presence of cholesterol oxidase?ChOx?can produce H₂O₂,the rGO/AuNP/luminol hybrid-based CL assay sensing platform could be used for cholesterol quantitation in the linear range from 0.71 to 11.43?M with a detection limit of 0.55?M.Besides,the CL sensor also can applied to assay other molecules by introducing other enzymes and substrates.?2?Graphitic carbon nitride nanosheet?g-C₃N₄ NS?was firstly found for effectively quenching the CL emission of luminol-H₂O₂ system due to the photoinduced electron transfer from luminol to g-C₃N₄ NS.Hence,a novel,label-free and versatile CL aptasensing platform was designed for sensitive detecting of mutiplue biomarkers,carcinoembryonic antigen?CEA?,adenosine triphosphate?ATP?and hepatitis B virus?HBV?by using aptamer as a recognition element.The G-rich DNA sequence of the DNA probe can self-assemble with hemin firstly to form hemin/G-quadruplex structure,a peroxidase-mimicking DNAzyme,which could catalyze luminol-H₂O₂ reaction to generate strong CL emission.When the target was added,a DNA-DNA duplex structure was formed owing to DNA hybridization reaction and target recognition effect,which could not be adsorbed onto the g-C₃N₄ NS surface because of its weak affinity.Thus,the electron transfer was blocked and the CL emission of luminol could be enhanced.There is a linear relationship between the CL intensity ratio?I/I₀?and CEA concentration?c?in the range of 0.1 ng/mL–150 ng/mL with a detection limit of63 pg/mL.The assay without any labelling or modifying,is facile and cost-effective,and will open a new avenue for the application of g-C₃N₄ NS in CL analysis.?3?A facile and sensitive chemiluminescence energy resonance transfer?CRET?aptasensor based on hemin/G-quadruplex DNAzyme was developed for miRNA-141detection.In this study,The hemin/G-quadruplex DNAzyme-catalyzed generation of CL through the oxidation of luminol by H₂O₂ stimulated the CRET to GO and fluorescein.The fluorescein-labeled DNA probes?G1,G1?binded with hemin to form a weak catalyst hemin/G-quadruplex DNAzyme and further absorbed on the surface of GO,leading to CL quenching of luminol.Upon the addition of miRNA-141,a stable and super catalyst hemin/G-quadruplex DNAzyme was formed and it could not absorbed onto the GO,weakening the CRET from luminol to GO.Therefore,the CL emission of luminol could be remarkably enhanced and the degree of CL increase is depended on the concentration of miRNA-141.There is a linear relationship between the CL intensity ratio?I/I₀?and miRNA-141 concentration?c?in the range of 0.01nM–200 nM with a detection limit of 4.7 pM.Additionally,the sensitivity of this method has been improved compared with that without fluorescein.The method can be extended to various detection by employing other DNA probes.In conclusion,we have fabricated a rGO hybrids with long-persistent CL,and developed novel luminol CL bioassays based on g-C₃N₄ NS and GO for the detection of biomolecule,protein,DNA,miRNA and small molecules.The proposed approaches may be extended to other target detection by simply changing the corresponding probe,which showing great promise for biomedicine and clinic diagnosis.Moreover,the research will open a new avenue for the application of graphene analogues nanomaterials in CL analysis,and conducive to expand application prospect of CL.