The Investigation Of Detecting ATP Based On Novel Carbon Nano-materials

Adenosine triphosphate(ATP), which exists in all organisms, plays an important and irreplaceable role in them, including: energy storage, energy transfer, synthesis of nutrients, promotion of cell repair and regeneration, maintaining the normal function of organisms. By detecting the ATP content, we can make an objective evaluations about killing effect of a drug or biological agents on cells, timely diagnosis and treatment some disease. In this study, two aptasensors based on novel carbon materials was developed for the rapid detection of ATP. Major works are as following:1. In this study, we have developed a biosensor to detect adenosine triphosphate(ATP), based on fluorescence resonance energy transfer(FRET) and making use of the activities of exonuclease I(EXO I) and exonuclease III(EXO III). In the absence of ATP in the biosensor reaction system, the aptasensor is hydrolyzed by EXO I. When ATP is present, it conjugates with the aptasensor and protects it from hydrolysis by EXO I; the aptasensor can then hybridize with a fluorescent sequence linked to graphene oxide(GO). The dsDNA formed by the interaction between the aptasensor and the fluorescent sequence is then recognized and cleaved by EXO III. The increased distance between the fluorescent particle(FAM, 6-carboxyfluorescein) and the GO significantly hinders the FRET and increases the fluorescence of FAM. By incorporating EXOIII into the process, the fluorescence signals of the biosensor are therefore greatly amplified and they were found to displayed a good linear relationship with ATP concentration, in the range from 0 to 3M. Moreover, this system can be employed for the detection of ATP in the real sample.2. Detection of ATP based on T-Hg2+-T complexes and aptamer. Carbon nanoparticles(CNPs), which can be quenched by mercury ion, were prepared by hydrothermal method. The existence of mercury iontriggered the formation of T-Hg2+-T complexes. In the absence of ATP, double-stranded structure containing T-Hg2+-T complexes can be stably existed in the solution, and CNPs existed in a free state, strong fluorescence can be detected. When ATP is present, it conjugates with the aptasensor quickly, resulted in the dissociation of double-stranded structure, and T-Hg2+-T complexes was destroyed, mercury ion is released from the double-stranded structure. Subsequently, the released mercury ion conjugates with CNPs which was in a free state. Because of the quenching effect of mercury ion, the fluorescence intensity of CNPs is weakened. With the increasing of ATP concentration, more and more Hg2 + is released, as a result, the fluorescence intensity of CNPs continues to weak until reached a plateau. Moreover, this system can be widely employed for the detection of other biological molecules when replacing the specific sequence of aptamer.