Aptamers as analytical reagents

Clark, Stacey L.; Remcho, Vincent T.

doi:10.1002/1522-2683(200205)23:9<1335::aid-elps1335>3.0.co;2-e

Cited by 145 publications

(74 citation statements)

References 31 publications

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“…This property has been exploited to create shape libraries that have yielded ligands that bind with high affinity and specificity to a variety of molecules across a broad range of size and chemical composition (29,30). A number of novel diagnostics and therapeutics have been isolated from these aptamer libraries, further indicating the significant amount of shape diversity they possess (31)(32)(33)(34)(35)(36). We have initiated studies to establish whether there is enough shape diversity in a nucleic acid aptamer library to allow recognition of each component of a complex biological mixture.…”

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confidence: 99%

Deconvolution of a Complex Target Using DNA Aptamers

Fitter¹,

James

2005

Journal of Biological Chemistry

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In vitro selection of single-stranded nucleic acid aptamers from large random sequence libraries is now a straightforward process particularly when screening with a single target molecule. These libraries contain considerable shape diversity as evident by the successful isolation of aptamers that bind with high affinity and specificity to chemically diverse targets. We propose that aptamer libraries contain sufficient shape diversity to allow deconvolution of a complex mixture of targets. Using unfractionated human plasma as our experimental model, we aim to develop methods to obtain aptamers against as many proteins as possible. To begin, it is critical that we understand how aptamer populations change with increasing rounds of in vitro selection when using complex mixtures. Our results show that sequence representation in the selected population changes dramatically with increasing rounds of selection. Certain aptamer families were apparent after only three selection rounds. Two additional cycles saw a decline in the relative abundance of these families and the emergence of yet another family that accounted for more than 60% of sequences in the pool. To overcome this population convergence, an aptamer-based target depletion method was developed, and the library screen was repeated. The previous dominant family effectively disappeared from the selected populations but was replaced by other aptamer families. Insights gained from these initial experiments are now being applied in the creation of second generation plasma protein screens and also to the analysis of other complex biological targets.

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confidence: 99%

Deconvolution of a Complex Target Using DNA Aptamers

Fitter¹,

James

2005

Journal of Biological Chemistry

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“…The three structures investigated are DNA aptamers [10,11] that were selected by the SELEX [10] method for their binding to the ligand L-argininamide (L-Arm) [12][13][14][15][16][17][18]. DNA and RNA aptamers are interesting molecules for two main reasons.…”

Section: Figure 1 Structure Of Ethidiummentioning

confidence: 99%

“…First, they comprise model systems for investigating the thermodynamic linkage between ligand binding and nucleic acid folding into bound complexes. Second, aptamers serve as potential novel therapeutics, catalysts, and biosensors based on their ability to bind a variety of ligands with affinities and selectivities rivaling monoclonal antibodies [11,[17][18][19]. Table 1 presents the sequence, Protein Data Bank (PDB) accession code, and length of the three aptamers studied here.…”

Section: Figure 1 Structure Of Ethidiummentioning

confidence: 99%

Probing the Structure of DNA Aptamers with a Classic Heterocycle.

Bishop

2004

Molecules

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DNA aptamers are synthetic, single-stranded DNA oligonucleotides selected by SELEX methods for their binding with specific ligands. Here we present ethidium binding results for three related DNA aptamers (PDB code: 1OLD, 1DB6, and 2ARG) that bind L-argininamide (L-Arm). The ligand bound form of each aptamer's structure has been reported and each are found to be composed primarily of two domains consisting of a stem helical region and a loop domain that forms a binding pocket for the cognate ligand. Previous thermodynamic experiments demonstrated that the DNA aptamer 1OLD undergoes a large conformational ordering upon binding to L-Arm. Here we extend those linkage binding studies by examining the binding of the heterocyclic intercalator ethidium to each of the three aptamers by fluorescence and absorption spectrophotometric titrations. Our results reveal that ethidium binds to each aptamer with ∆G o 's in the range of -8.7 to -9.4 kcal/mol. The stoichiometry of binding is 2:1 for each aptamer and is quantitatively diminished in the presence of L-Arm as is the overall fluorescence intensity of ethidium. Together, these results demonstrate that a portion of the bound ethidium is excluded from the aptamer in the presence of a saturating amount of L-Arm. These results demonstrate the utility of ethidium and related compounds for the probing of non-conventional DNA structures and reveal an interesting fundamental thermodynamic linkage in DNA aptamers. Results are discussed in the context of the thermodynamic stability and structure of each of the aptamers examined.Molecules 2004, 9 68

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“…The development of aptamer technology significantly broadens the applications of nucleic acids. They are widely used as recognition element and applied in capillary electrophoresis [16], affinity chromatography [17], and biosensors [18,19]. Comparing with antibody, aptamers are conveniently and economically synthesized, flexibly chemical-modified, stable and less vulnerable to denaturation.…”

Section: Introductionmentioning

confidence: 99%

A novel separation and enrichment method of 17β-estradiol using aptamer-anchored microbeads

Huy

Jin

Yin

et al. 2010

Bioprocess Biosyst Eng

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The estrogenic compound 17b-estradiol (E2) is widely studied for its potential endocrine disruption effects. Due to the low level of E2 present in the environment, it is highly desirable to develop a sensitive and efficient separation and enrichment method for E2 analysis. In this paper, we proposed a novel E2 preconcentration method using anti-E2 aptamer-anchored isothiocyanate-modified beads (NCS beads). The glass beads are chemically modified with primary amino group, and then treated with phenylene diisothiocyanate (PDITC) to generate an isothiocyanate group, which is reactive towards the amine group. The amino-modified anti-E2 aptamer can be easily covalently immobilized onto the as-prepared NCS beads. The experimental results demonstrated that the aptamer affinity microbeads could selectively retain and separate E2 compound. The effects of the operation parameters on retention of E2, including washing condition, eluting condition, the number of beads, and the incubation time were investigated. Moreover, high-performance liquid chromatography with preconcentration of E2 on the aptamer affinity microbeads was applied to detect the E2 in the spiked water samples and obtained a good recovery.

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Aptamers as analytical reagents

Cited by 145 publications

References 31 publications

Deconvolution of a Complex Target Using DNA Aptamers

Deconvolution of a Complex Target Using DNA Aptamers

Probing the Structure of DNA Aptamers with a Classic Heterocycle.

A novel separation and enrichment method of 17β-estradiol using aptamer-anchored microbeads

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