Competitive Assay for Theophylline Based on an Abasic Site-Containing DNA Duplex Aptamer and a Fluorescent Ligand

A fluorescence assay for theophylline, one of the common drugs for acute and chronic asthmatic conditions, has been developed based on an abasic site‐containing DNA duplex aptamer (AP aptamer) in combination with an abasic site‐binding fluorescent ligand, riboflavin. The assay is based on the compet...

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Bibliographic Details
Published in:Chemistry : a European journal 2012-10, Vol.18 (40), p.12719-12724
Main Authors: Sato, Yusuke, Zhang, Yushuang, Nishizawa, Seiichi, Seino, Takehiro, Nakamura, Kodai, Li, Minjie, Teramae, Norio
Format: Article
Language:English
Subjects:
abasic sites
aptamers
Aptamers, Nucleotide - chemistry
Assaying
Binding
Binding Sites
Chemistry
Competition
Cytosine - chemistry
Deoxyribonucleic acid
DNA
DNA - chemistry
DNA binding
Fluorescence
fluorescent probes
Humans
Ligands
Molecular Sequence Data
Nucleotides - chemistry
Polymorphism, Single Nucleotide
Receptors
Riboflavin
Riboflavin - analysis
Riboflavin - chemistry
Spectrometry, Fluorescence
Theophylline
Theophylline - analysis
Theophylline - blood
Theophylline - chemistry
Vitamin B
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Summary:A fluorescence assay for theophylline, one of the common drugs for acute and chronic asthmatic conditions, has been developed based on an abasic site‐containing DNA duplex aptamer (AP aptamer) in combination with an abasic site‐binding fluorescent ligand, riboflavin. The assay is based on the competitive binding of theophylline and riboflavin at the abasic (AP) site of the AP aptamer. In the absence of theophylline, riboflavin binds to the receptor nucleotide opposite the AP site, which leads to fluorescence quenching of the riboflavin. Upon addition of theophylline, competitive binding occurs between theophylline and riboflavin, which results in an effective fluorescence restoration due to release of riboflavin from the AP site. From an examination of the optimization of the AP aptamers, the complex of riboflavin with a 23‐mer AP aptamer (5′‐TCT GCG TCC AGX GCA ACG CAC AC‐3′/5′‐GTG TGC GTT GCC CTG GAC GCA GA‐3′; X: the AP site (Spacer C3, a propylene residue)) possessing cytosine as a receptor nucleotide was found to show a selective and effective fluorescence response to theophylline; the limit of detection for theophylline was 1.1 μM. Furthermore, fluorescence detection of theophylline was successfully demonstrated with high selectivity in serum samples by using the optimized AP aptamer and riboflavin. Lighting up the competition: A fluorescence assay is reported for the detection of theophylline by using an abasic site (AP site)‐containing DNA duplex aptamer (AP aptamer) in combination with an abasic site‐binding fluorescent ligand, namely riboflavin. The assay is based on the competitive binding of theophylline or riboflavin at the AP site of the AP aptamer (see scheme). Fluorescence detection of theophylline was successfully demonstrated with high selectivity in serum samples by using the optimized AP aptamer and riboflavin.
ISSN:0947-6539
1521-3765
DOI:10.1002/chem.201201298