Amperometric Biosensing of miRNA-21 in Serum and Cancer Cells at Nanostructured Platforms Using Anti-DNA–RNA Hybrid Antibodies

Abstract

This paper describes a disposable amperometric biosensor for simple and sensitive determination of miRNA-21. The bioplatform consists of gold nanoparticles-nanostructured electrode surfaces on which a direct hybridization assay involving the immobilization of a specific thiolated DNA capture probe and recognition of the formed DNA–miRNA-21 heteroduplex by a specific antibody is implemented. The antibody is further recognized through its Fc region by a commercial bacterial protein containing 40 units of horseradish peroxidase (HRP) (ProtA-polyHRP40). The amperometric detection of the hybridization process using the H 2 O 2 /hydroquinone system allows quantification of the target miRNA in the 0.096–25 pM linear range with a detection limit of 29 fM (0.29 amol in 10 μL of sample). The bioplatform offers excellent selectivity against noncomplementary sequences and very acceptable against sequences with an unpaired base (only 30% of the response obtained for the target miRNA). In addition, the bioplatform was shown to be useful for the determination of the endogenous content of the target oncomiR directly in blood serum from breast cancer patients and in breast cancer cells using only 10 ng of total extracted RNA.

A miRNA biosensor based on localized surface plasmon resonance enhanced by surface-bound hybridization chain reaction

“…2 , LOD and LOQ are the lower and higher horizontal dashed lines, for both HCR-less and HCR-enhanced measurements). These LOD and LOQ values are in the proper range for miRNA detection in bodily fluids, where the concentrations are expected to range from femtomolar to nanomolar ( Zouari et al, 2018 ). As the volume of each target analyte injection was 50 μL, the LSPR biosensor can detect about 50 amol of miRNA.…”

Advances in Electrochemical (Bio)Sensing Targeting Epigenetic Modifications of Nucleic Acids

“…Amperometric biosensing methods for the single (a) and multiplexed (b) miRNAs determination involving a direct DNA/ miRNA hybridization onto AuNPs-SPCE and labelling of the RNA heterohybrids with Ab DNA-RNA and ProtA-Poly-HRP40 (a), or an HCR-based immunosensing strategy using Ab DNA-RNAmodified MBs for capturing the target miRNAs. Reprinted from [54] (a) and [55] (b) with permission. Tavallaie et al [29] have proposed recently an attractive electrochemical sensor for the determination of miRNAs based on the use of an electrically reconfigurable DANN-gold-coated magnetic nanoparticles (Au@MNPs) network (DNA-Au@MNPs).…”

“…Among the methods using bioreceptors with a high affinity towards RNA hybrids, strategies involving the Ab DNA‐RNA antibody or the viral protein p19 for the recognition of hetero‐ and homo‐ RNA hybrids, respectively, were implemented on the surface of Strep‐MBs or AuNPs‐SPCEs, in connection with biotinylated and thiolated Cp, respectively. These methodologies leveraged the selectivity of target miRNA/complementary probe hybridization with the unique specificity of the bioreceptors for particular RNA duplexes.…”

“…These methodologies leveraged the selectivity of target miRNA/complementary probe hybridization with the unique specificity of the bioreceptors for particular RNA duplexes. In addition, they were smartly used together with amplification strategies of easy implementation in POC devices such as ProtA‐Poly‐HRP40 multienzyme labels (Figure a) or HCR (Figure b), which led to excellent sensitivity, particularly in the case when nanostructured platforms were used, providing LOD values of 29 and 142 fM for the amperometric detection of the synthetic target miRNA. This improved sensitivity was attributed to the enlarged active surface area, the inherent conductivity and catalytic activity of AuNPs as well as to the efficient immobilization of the thiolated nucleic acid probe with an adequate orientation and spacing thus improving the target accessibility.…”

Biosensing and Delivery of Nucleic Acids Involving Selected Well-Known and Rising Star Functional Nanomaterials

“…The use of AuNPs as electrode modifiers provides huge active surface area and facilitates the electron transfer processes at the sensing interface, thereby granting a high sensitivity to biosensors. In addition to the improved conductivity and the catalytic activity of AuNPs, nanostructured surfaces with these nanoparticles greatly enhance the amount of immobilized thiolated probes and allow their favorable orientation and spacing, thus ensuring the target accessibility and improving the hybridization efficiency [2,8,9,10,11]. The great biocompatibility of AuNPs is essential to keep the biological activity of the biomolecules attached to their surface, which produces the attractive storage stability of the prepared bioplatforms [10,11].…”