The Research Of Immunoglobulin E Detection Method Based On Fluorescent Sensor

Immunoglobulin E(IgE), as a secretory immunoglobulin, plays a key role in human allergic responses. Normally, IgE is present in small amouts in human blood serum, however, IgE levels increase in patients with allergic asthma, atopic dermatitis and other immune deficiency-related diseases. Therefore, it’s important for clinical diagnostics to develope a simple, rapid and sensitive method for the detection of IgE. Combining fluorescence spectroscopy and aptamer possessing high specificity, this article established a fluorescent aptasensor based on the principle of fluorescence resonance energy transfer(FRET) to detect immunoglobulin E (IgE).In this article, Cyanine dyes — Cy5 and hydrophilic carbon nanomaterials — graphene oxide (GO) were chosen as the fluorescent energy donor and receptor respectively. The anti-IgE aptamer whose tagged with Cy5 could be steadily absorbed on the surface of GO via π-π conjugate accumulation effect. Resonance energy transfer between Cy5 and GO. could quench the fluorescence of Cy5. With the target protein IgE added, it competed with GO for binding to aptamer. Thus, the fluorescent signals of Cy5 restored. In the experimental process, we firstly used the Three-Dimensional Excitation Emission matrix Fluorescence Spectrophotometer (3D-EMM) to analyze the properties of Cy5 tagged aptamer, using 633 nm and 675 nm as the excitation and emission wavelength, respectively. The kind of quenchers, incubation temperature and incubation time were optimized. The results showed that the highest fluorescence signal response was obtained using GO as the quencher after incubation in 1×PBS(pH 7.4, CMg2+1 mmol/L) at 37℃ for 30 min. Under the optimized conditions, linear range of 0.05-10 nmol/L was obtained, with a detection limit of 1 pmol/L. IgG and HAS were selected for the specificity validation of the present method. The result indicated that, both showed low fluorescence response, which indicates a high selectivity and specificity of this method.Finally, we applied the established method to detect the IgE in a human blood sample, and used the classical ELISA method to verify the test result, which exhibited a good consistency. We conclude that the present method is efficient, rapid, sensitive and has great potential in actual clinical applications.