Aptamer-based Piezoelectric Quartz Crystal Microbalance Biosensor Array For The Quantification Of IgE And Comparison With The Piezoelectric QCM Immunosensor

Objectives:The aim of this study was to develop a rapid method to measure IgE in human serum by use of a direct aptamer-based biosensor based on a quartz crystal microbalance (QCM). The comparison with the antibody-based QCM immunosensor was also explored.Methods:1. We designed and fabricated aptamer-based piezoelectric quartz crystal microbalance biosensor. Self-oscillating TTL circuit and self-made optimized PESA version 4.0 software applied to record and analyze frequency changes.2. An avidin monolayer prepared by the covalently method was applied to immobilize biotinylated single-stranded DNA aptamers specific for IgE on the gold surface of the quartz crystal. The frequency shifts (FS) of the QCM were measured and related to IgE concentrations. The dose-curve was drawn according to the relationship betweenΔF and IgE concentrations.3. The sensitivity, detection limit, nonspecific response and regeneration ability of piezoelectric QCM aptamer biosensor were observed. At last, clinical samples were detected in order to explore the possibility about the detection of clinical samples. The aptamer piezoelectric biosensor used no labeled reagent and no pretreatment of samples.4. The sensitivity, detection limit, nonspecific response and regeneration ability of the QCM aptamer biosensor also compared with the antibody-based QCM immunosensor.Results:1. A linear relationship existed between the FS (Hz) and the IgE concentrations from2.5 to 200μg/L in buffer and human serum. The correlation coefficient between QCM method and chemiluminescence method was 0.992 for determination of IgE in 50 clinical human serum samples.2. The cross-reactivity of the aptamer was seen to be less than 5% for all the interference proteins.3. The aptamer receptors tolerated repeated affine layer regeneration after ligand binding and recycling of the biosensor with little loss of sensitivity. Using 30 mmol/L EDTA as regeneration reagent gave the best result. More than 80% of the original response was maintained after 10 assays.4. The stability of the aptamer biosensor was almost fully maintained at the first 7 days and then started to decline. When stored for 3 weeks, the FS were all >90% of those on the response at the first day.5. The aptamer biosensor and QCM immunosensor were able to detect IgE with high specificity and sensitivity in 15 min. The results of them were compared with chemiluminescence method, the correlation coefficients were good.6. The regeneration ability and long-term stability of QCM aptamer biosensor were better than that of piezoelectric QCM immunosensor.Conclusions:1. The correlation coefficient between QCM aptamer biosensor and chemiluminescence was 0.992 for determination of IgE in the clinical human serum samples. The results showed that the aptamers was able to detect IgE with high sensitivity and selectivity, even in a complex biological sample such as serum.2. The QCM aptamer biosensor and QCM immunosensor capable of detecting IgE within 15 minutes, provides patients with a faster diagnosis due to a shorter detection time, and the use of relatively easy-to-use devices. Moreover, the biosensor method has shown its advantages over other conventional methods due to its label-free property. The direct biosensor system measured IgE without the need of a labeled reagent and pretreatment of samples.3. The precision, detection limit, specificity and accuracy of the aptamer biosensor were better than that of immunosensor.4. The reusability and long-term stability of the aptamer biosensor were well, which can reduce the cost of detection.5. The QCM biosensor can measure IgE and offer advantages of high specificity, reusability, low detection limit, no label or sample pretreatment, and shorter detection time. The aptamer QCM biosensor was suitable for sensitive and specific protein detection, representing an innovative tool for future proteomics.