Studies On Fluorescence Aptasensor And Application In Detection Of Metal Ions And Biological Small Molecules

With the development of society and the improvement of people’s living standard, environmental problems and people’s health have attracted more and more attentions. It is significant to analyze and detect metal ions and biological small molecules in the area of the prevention and diagnosis of disease and the improvement of people life quality. The appearance of aptamer brings new power for the area of biochemical analysis. Aptamers are artificial single-stranded oligonucleic acids selected in vitro, and they have high affinity and selectivity towards a variety of targets, including small molecules, proteins, metal ions, drugs, and even whole cells. Aptasensors become new emerging forces in analysis field because of the advantages of aptamers such as high selectivity, extensive target, easier synthesis and modification, good stability and so on. In this paper, we have developed three kinds of fluorescence aptasensors for the detection of metal ions and biological small molecules by combining aptamer with fluorescent technology. The main contents are summarized as follows:1. A novel label-free fluorescent aptasensor for the detection of potassium ion (K+) was developed based on the horseradish peroxidase-mimicking DNAzyme (HRP-DNAzyme). In the presence of K+, the G-rich DNA folded into the G-quadruplex structure, and then hemin can bind to the G-quadruplex structure as a co-factor and form HRP-DNAzyme.3-(p-Hydroxyphenyl)-propanoic acid (HPPA) can be oxidized by H2O2into a fluorescent product in the presence of DNAzyme. The fluorescence intensity of the HPPA oxidative product increased with the K+concentration. Under the optimal conditions, the fluorescence intensity was linearly related to the logarithm of K+concentration in the range of2.5μM to5mM. Other metal ions caused no notable interference on the detection of K+. The proposed fluorescence aptasensor is simple, sensitive and selective, it shows widely promising application in ions detection and disease diagnosis.2. A label-free and sensitive OFF-ON-OFF fluorescent aptasensor was constructed for the detection of Ag+and dopamine based on the formation of C-Ag+-C conjugates. In the presence of Ag+, the Ag+binds directly to N3of cytosine to form metal mediated C-Ag+-C duplexes, and then SG can integrate with duplexes structure and yield high fluorescence. The fluorescence intensity of SG increased with the Ag+concentration, leading to a development of fluorescent "turn on" sensor for Ag+ions. Addition of dopamine causes fluorescence signal decrease, the fluorescence intensity of SG decreased with the increasing in the concentration of dopamine concentration, leading to a development of fluorescent "turn off’ sensor for dopamine. The proposed sensor is simple, sensitive and it could detect Ag+and dopamine down to concentration of3.3nM and1μM, respectively, opening a new method for the development of dual-output detection.3. Based on the guanine (G)-quenching, a simple and sensitive fluorescence aptasensor for the detection of adenosine was constructed. The FAM-labeled adenosine aptamer is partly complementary to DNA sequence containing five G in its end, in the absence of adenosine, the hybridization of FAM-labeled aptamer with complementary strand cause G approach to FAM, hence, the fluorescence of FAM was effectively quenched because of the photoinduced electron transfer mechanism (PET) between FAM and G. Upon addition of adenosine, it reacts with FAM-labeled aptamer, allowing complementary strand to release and fluorescence intensity to restore. The linear range of proposed adenosine sensor is20-200μM, and detection limit down to6.7μM (S/N=3). Most importantly, it don’t need to use organic dye molecules-labeled DNA or extra nanomaterials as quenchers, it is simple, sensitive and low-cost.