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Detection Methods of Adenosine Based on Aptamer and Their Application

Author: ZhangJinQuan
Tutor: WangYongSheng
School: Nanhua University
Course: Health Toxicology
Keywords: Adenosine Aptamer Gold nanoparticles Resonance light scattering G-quartet Fluorescence resonance energy transfer Aptamer structure switch
CLC: R115
Type: Master's thesis
Year: 2010
Downloads: 78
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Abstract

Adenosine (AD) is an endogenous nucleoside which has received much attention due to its crucial signaling functions in both the peripheral and central nervous system. A number of investigations strongly suggest that solid tumors routinely experience severe hypoxia and necrosis owing to their rapid growth, resulting in degradation of adenine nucleotide to release adenosine. The released adenosine would benefit malignancy by providing a supportive environment for the accelerated growth, and lead to accumulation of adenosine of high concentration, which has been demonstrated in solid tumors. Consequently, the concentration of urinary adenosine reflects level of adenine nucleotide degradation in the organism. Some of studies have examined its biomedical significance as possible biomarkers for cancer. Urinary adenosine also has been used to monitor progress of diseases. In addition, direct monitoring of adenosine fluctuations under physiological conditions would also play an important role in the investigation related to heart and brain physiology.In the chapter 2, a novel sensitive method has been developed for the detection of adenosine (AD) in human urine by using enhanced resonance light scattering (RLS). This method is based on the specific recognition and signal amplification of adenosine aptamer (Apt) coupled with gold nanoparticles (GNPs) via G-quartet induced nanoparticle assembly, which was fabricated by triggering a structure-switching of 3’terminus G-rich sequence and aptamer duplex. RLS signal linearly correlated with the concentration of adenosine over the range of 6.0×10-9～11.5×10-8 mol·L-1. The limit of detection (LOD) for adenosine is 1.8×10-9 mol·L-1 with relative standard deviations (R.S.D) of 3.55%, 4.37%, 5.85%, (n = 6) and the recoveries were 96.3%, 98.8%, 103.4% (n=6), respectively. The present method has been successfully applied to determination of adenosine in real human urine, and the obtained results were in good agreement with those obtained by the HPLC method. Our investigation shows that the combination of the excellent selectivity of aptamer with high sensitivity of RLS technique could provide a promising potential for aptamer-based small molecules detection, and be benefit to extend the application of RLS.In the chapter 3, a new method for the detection of adenosine (AD) in human urine has been established by using structure-switching aptamer coupled with 6-FAM and gold nanoparticles as fluorescence resonance energy transfer donor and recepter, respectively. The measurement was carried out in 20 mmol·L-1 Tris-HCl buffer solution of pH 7.4,ΔF signal linearly correlated with the concentration of adenosine over the range of 2.0×10-8～1.8×10-6 mol·L-1. The limit of detection (LOD) for adenosine is 6.0×10-9mol·L-1. The present method has been successfully applied for the determination of adenosine in artificial human urine samples. The relative standard deviations were 5.25%, 3.64%, 5.36%, and the recoveries were 97.6%, 102.1%, 105.8% (n=6), respectively. The present method has been successfully applied to determination of adenosine in real human urine samples, and the obtained results were in good agreement with those obtained by the HPLC method.

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