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The Application of Microfluidic Technologies in Aptamer Selection.

ABSTRACT: Aptamers are sequences of single-strand oligonucleotides (DNA or RNA) with potential binding capability to specific target molecules, which are increasingly used as agents for analysis, diagnosis, and medical treatment. Aptamers are generated by a selection method named systematic evolution of ligands by exponential enrichment (SELEX). Numerous SELEX methods have been developed for aptamer selections. However, the conventional SELEX methods still suffer from high labor intensity, low operation efficiency, and low success rate. Thus, the applications of aptamer with desired properties are limited. With their advantages of low cost, high speed, and upgraded extent of automation, microfluidic technologies have become promising tools for rapid and high throughput aptamer selection. This paper reviews current progresses of such microfluidic systems for aptamer selection. Comparisons of selection performances with discussions on principles, structure, operations, as well as advantages and limitations of various microfluidic-based aptamer selection methods are provided.

SUBMITTER: Liu Y

PROVIDER: S-EPMC8484746 | biostudies-literature | 2021

REPOSITORIES: biostudies-literature

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The Application of Microfluidic Technologies in Aptamer Selection.

Liu Yang Y Wang Nijia N Chan Chiu-Wing CW Lu Aiping A Yu Yuanyuan Y Zhang Ge G Ren Kangning K

Frontiers in cell and developmental biology 20210917

Aptamers are sequences of single-strand oligonucleotides (DNA or RNA) with potential binding capability to specific target molecules, which are increasingly used as agents for analysis, diagnosis, and medical treatment. Aptamers are generated by a selection method named systematic evolution of ligands by exponential enrichment (SELEX). Numerous SELEX methods have been developed for aptamer selections. However, the conventional SELEX methods still suffer from high labor intensity, low operation e ...[more]

PMID: 34604229

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Project description:Brevetoxins (PbTxs) are very potent marine neurotoxins that can cause an illness clinically described as neurologic shellfish poisoning (NSP). These toxins are cyclic polyether in chemistry and have increased their geographical distribution in the past 2 decades. However, the ethical problems as well as technical difficulties associated with currently employed analysis methods for marine toxins have spurred the quest for suitable alternatives to be applied in a regulatory monitoring regime. In this work, we reported the first instance of concurrent aptamer selection of Brevetoxin-1 (PbTx-1) and Brevetoxin-2 (PbTx-2) and constructed a biolayer interferometry (BLI) biosensor utilizing PbTx-1 aptamer as a specific recognition element. Through an in vitro selection process, we have, for the first time, successfully selected DNA aptamers with high affinity and specificity to PbTx-1 and PbTx-2 from a vast pool of random sequences. Among the selected aptamers, aptamer A5 exhibited the strongest binding affinity to PbTx-1, with an equilibrium dissociation constant (KD) of 2.56 μM. Subsequently, we optimized aptamer A5 by truncation to obtain the core sequence (A5-S3). Further refinement was achieved through mutations based on the predictions of a QGRS mapper, resulting in aptamer A5-S3G, which showed a significant increase in the KD value by approximately 100-fold. Utilizing aptamer A5-S3G, we fabricated a label-free, real-time optical BLI aptasensor for the detection of PbTx-1. This aptasensor displayed a broad detection range from 100 nM to 4000 nM PbTx-1, with a linear range between 100 nM and 2000 nM, and a limit of detection (LOD) as low as 4.5 nM. Importantly, the aptasensor showed no cross-reactivity to PbTx-2 or other marine toxins, indicating a high level of specificity for PbTx-1. Moreover, the aptasensor exhibited excellent reproducibility and stability when applied for the detection of PbTx-1 in spiked shellfish samples. We strongly believe that this innovative aptasensor offers a promising alternative to traditional immunological methods for the specific and reliable detection of PbTx-1.

Dataset Information

The Application of Microfluidic Technologies in Aptamer Selection.

Publications

The Application of Microfluidic Technologies in Aptamer Selection.

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