Concatemeric DNA Research Articles

High-Plex ELISPOT: FOLISPOT Based on Fluorescence Detection and DNA Complementary Pairing.

Enzyme-linked immunospot (ELISPOT) is one of the most important methods to measure the number of specific cells by detecting protein secretion at a single-cell level. However, traditional ELISPOT based on enzyme-substrate color development can only detect one target. Therefore, scientists developed multiple-target ELISPOT based on enzyme-substrate coloring. Besides, FluoroSPOT that can detect 2-4 fluorescent signals are developed. Nevertheless, the maximum detection targets of multiple-target ELISPOT and FluoroSPOT are around 4, and the signal amplification system can be further optimized. Fluorescence-based oligo-linked immunospot (FOLISPOT), which utilized DNA-barcoded antibodies to provide a highly multiplexed method with signal amplification, was developed to detect multiple targets simultaneously. In this method, multiple targets can be detected in one round and multiple rounds of detection can be conducted, and thus a large number of targets can be detected. Besides, signal amplification is achieved by DNA complementary pairing and modular orthogonal DNA concatemers, and thus cells secreting limited amounts of proteins can be detected. According to the studies, FOLISPOT can detect more spots than ELISPOT and can detect targets that are undetectable by ELISPOT. Furthermore, FOLISPOT can be utilized to detect more than 6 targets, by allowing sequential detection of multiple targets in one round and sequential detection in multiple rounds.

Identification of a leucine-zipper motif in pUL51 essential for HCMV replication and potential target for antiviral development

Human cytomegalovirus (HCMV) can cause serious diseases in immunocompromised patients. Use of current antivirals is limited by their adverse effects and emergence of drug resistance mutations. Thus, new drugs are an urgent need. The terminase complex (pUL56-pUL89-pUL51) represents a target of choice for new antivirals development. pUL51 was shown to be crucial for the cleavage of concatemeric HCMV DNA and viral replication. Its C-terminal part plays a critical role for the terminase complex assembly. However, no interaction domain is clearly identified. Sequence comparison of herpesvirus homologs and protein modelling were performed on pUL51. Importance of a putative interaction domain is validated by the generation of recombinant viruses with specific alanine substitutions of amino acids implicated in the domain. We identified a Leucine-Zipper (LZ) domain involving the leucine residues L126-X6-L133-X6-L140-X6-L147 in C-terminal part of pUL51. These leucines are crucial for viral replication, suggesting the significance for pUL51 structure and function. A mimetic-peptide approach has been used and tested in antiviral assays to validate the interaction domain as a new therapeutic target. Cytotoxicity was evaluated by LDH release measurement. The peptide TAT-HK29, homologous to the pUL51-LZ domain, inhibits HCMV replication by 27% ± 9% at 1.25 μM concentration without cytotoxicity. Our results highlight the importance of a leucine zipper domain in the C-terminal part of pUL51 involving leucines L126, L133, L140 and L147. We also confirm the potential of mimetic peptides to inhibit HCMV replication and the importance to target interaction domains to develop antiviral agents.

An electrochemical aptasensor for exosomes based on strand displacement amplification and hybridization chain reaction amplification

Exosomes are lipid bilayer vesicles, relating to the happening and progress of cancer. However, exploring reliable, convenient and sensitive exosomes detection methods in early cancer diagnosis is still challenging. Here, an electrochemical aptasensor based on both signal amplifications of strand displacement amplification (SDA) and hybridization chain reaction (HCR) was developed to sensitively detect tumor exosomes. Firstly, exosomes were recognized by CD63 aptamer, and converted into DNA concentration by chain replacement reaction and magnetic separation. A produced DNA fragment (P) was obtained by SDA，then P was captured at the modified gold electrode and amplified using HCR. Abundant horseradish peroxidase (HRP) attached to DNA concatemers and catalyzed the oxidation of o-phenylenediamine (OPD). The reduction current of 2,3-diaminopenazine (DAP) was measured and the exosomes were quantized. Under the optimal conditions, the DPV response showed a great linear relationship with exosomes concentration range of 44 ∼ 8.8 × 106 particle/μL. The limit of detection was 18 particle/μL. Experimental results show that this is an alternative method to detect exosomes and other tumor markers.

Highly repeatable detection of miRNAs in complex biological mixtures mediated by magnetically separable spherical nucleic acids and homogeneous signal amplification

Immobilization-free photoelectrochemical (PEC) sensing holds a great potential in achieving tumor biomarker detection with high repeatability. However, most sensors suffer from weak signal and background interference, which severely limits its practical translational application toward the analysis of complex physiological samples. Herein, a homogeneous and label-free miRNA sensing system was developed by integrating magnetically separable spherical nucleic acids (SNA) and signal amplification in PEC detection. Firstly, target-capturing probes were covalently conjugated to the surface of polydopamine-coated Fe3O4 nanoparticles. SNA structures consisting of chain-extended DNA concatemers from analyte-initiated hybridization chain reactions (HCR) served as hosting reservoirs of photosensitizers (PS) to achieve signal conversion. Subsequently, magnetic separation was applied to remove complex biological matrix and PS intercalated SNA particles. The concentration signal of low abundance targets was reflected in the difference in PS concentration in the supernatant. The electron transfer on electrode surface was boosted by redox mediators in homogeneous PEC detection. Finally, the proposed nanosensing system achieved highly repeatable (relative standard deviation 2.5–3.1 %) and the anti-interference analysis of miR-21 (limit-of-detection, 0.33 fM). The practical applicability was illustrated in sensitive profiling of miR-21 in lysate of tumor cells and plasma of tumor-bearing mice.

Concentration of lambda concatemers using a 3D printed device.

Identifying significant variations in genomes can be cumbersome, as the variations span a multitude of base pairs and can make genome assembly difficult. However, large DNA molecules that span the variation aid in assembly. Due to the DNA molecule's large size, routine molecular biology techniques can break DNA. Therefore, a method is required to concentrate large DNA. A bis-acrylamide roadblock was cured in a proof-of-principle 3D printed device to concentrate DNA at the interface between the roadblock and solution. Lambda concatemer DNA was stained with YOYO-1 and loaded into the 3D printed device. A dynamic range of voltages and acrylamide concentrations were tested to determine how much DNA was concentrated and recovered. The fluorescence of the original solution and the concentrated solution was measured, the recovery was 37% of the original sample, and the volume decreased by a factor of 3 of the original volume.

Ultrasensitive electrochemical sensing platform for miRNA-21 detection based on manganese dioxide-gold nanoparticle nanoconjugates coupled with hybridization chain reaction and horseradish peroxidase signal amplification.

In this study, an ultrasensitive electrochemical miRNA-21 biosensor is described. Manganese dioxide-gold nanoparticle (MnO2-Au NP) nanoconjugates were employed as sensing base materials, miRNA-21 was selected as a model analyte, and hybridization chain reaction (HCR) was employed to form long DNA concatemers using two different oligonucleotides with a complementary sequence. Thus, lots of biotin were loaded on DNA concatemers and one of them was labelled with biotin at its 3' terminal. The biosensor was designed as follows: a sulfhydryl-hairpin probe (HP) was first dropped on the surface of the glassy carbon electrode (GCE) modified with MnO2-Au NP nanoconjugates (HP/MnO2-AuNPs/GCE). After it was treated with MCH, the modified electrode was hybridized with miRNA-21, resulting in the loop of HP being opened to form a vertical structure. Subsequently, the modified electrode (miRNA-21/HP/MCH/MnO2-AuNPs/GCE) was incubated with DNA concatemers to form a sandwich structure of HP-miRNA-21-DNA concatemers on the modified electrode surface. Finally, the streptavidin-HRP conjugates were linked to the sandwich structure by specific recognition interaction between biotin and avidin. Differential pulse voltammetry (DPV) was used to measure the electrochemical response of the biosensor in the phosphate-buffered solution (0.10 M PBS, pH 7.0) containing 2.0 mM hydroquinone (HQ) and 1.8 mM H2O2. As a result, a larger reductive signal was obtained at a potential of -0.17 V (vs. SCE). Various experimental conditions were optimized, including solution pH, incubation time, and the amount of DNA concatemers. Under optimal conditions, the biosensor showed good sensing performance, such as a wide linear response range (0.1 fM and 100 nM) and low detection limit (0.063 fM, at S/N = 3). Meanwhile, the biosensor can discriminate single base matched miRNA-21, indicating that the biosensor had good selectivity.

A methylene blue-functionalized DNA concatemer for the construction of a turn-off fluorescent immunosensor for the sensitive detection of carcinoembryonic antigen.

This study showed a method of synthetization of a methylene blue-functionalized DNA concatemer via hybridization chain reaction (HCR) used for turn-off fluorescence detection of carcinoembryonic antigen (CEA). During the experiments, CEA aptamers and the methylene blue-functionalized DNA concatemer were modified onto the surface of Au nanoparticles (AuNPs). By detecting the signal of remaining methylene blue in the solution that has not been embedded in the DNA concatemer, we obtained an amplified decrease of the fluorescence signal at 695 nm for CEA. The linear range was from 0.1 to 80 ng mL-1 with a limit of detection at 75 pg mL-1 for CEA determination. Our results showed that the proposed method had good selectivity and could generate satisfactory results for clinical serum sample determination. Based on the positive outcomes obtained, we determined that the method provided a sensitive and accurate way for early clinical diagnosis of cancer disease.

One lead to numerous: A DNA concatemer-based fluorescence aptasensor for selective and sensitive Acinetobacter baumannii detection

Rapid and precise analysis of Acinetobacter baumannii plays a crucial role in inhibition of healthcare-associated infectious diseases and development of optimal treatment strategies. Previously reported fluorescence sensors have been promising, but were hindered by poor selectivity and low sensitivity due to the complex detection environment and limited identification ability. Thus, specifically recognised and effectively amplified strategies are vital for overcoming the shortcomings of existing analytical methods. Herein, a highly sensitive and selective fluorescence aptasensor was constructed for the quantification of A. baumannii based on aptamer-mediated DNA concatemers. The self-assembled DNA concatemers were anchored on the surface of micromagnetic beads by aptamers as an intermediation embellished with numerous carbon quantum dots (CQDs). Effective recognition and capture of A. baumannii cells by the aptamer immediately triggered the release of DNA concatemers. Meanwhile, a magnified fluorescent signal wherein a single A. baumannii cell results in numerous CQDs would be generated. The as-prepared biosensor exhibited an excellent detection and quantification limit (LOD) as 52.67 (11.20) cells/mL, LOQ = 155.3 (10.80) cells/mL over a wide linear range (9.80×102 to 1.25×105 cells/mL) for A. baumannii analysis, which was superior to previously reported methods. Furthermore, the sensor exhibited satisfactory accuracy and good reproducibility in urine samples obtained from patients with bladder cancer and healthy individuals.

Tuning the Surface Molecular Charge of Organic Photoelectrochemical Transistors with Significantly Improved Signal Resolution: A General Strategy toward Sensitive Bioanalysis.

Nature makes use of molecular charges to operate specific biological synthesis and reactions. Targeting advanced opto-bioelectronic sensors, organic photoelectrochemical transistors (OPECTs), taking advantage of the light fuel substituting an external gate potential, is now debuting and expected to serve as a universal platform for studying the rich light-biomatter interplay for new bioanalytics. Given the ubiquity of charged biomolecules in nature, molecular charge manipulation should underpin a generic route for innovative OPECT regulation and operation, which nevertheless has remained unachieved. Herein, this work manifests the biological tuning of surface charge toward the OPECT biosensor, which was exemplified by a light-sensitive CdS quantum dot (QD) gate electrode interfaced by a smart DNA superstructure with adenosine triphosphate (ATP) responsiveness. Highly negative-charged supramolecular DNA concatemers were self-assembled via sequential hybridization, and the ATP-triggered disassembly of the DNA concatemers would cause a tandem change of the effective gate voltage and transfer characteristics with significantly improved resolution. The present opto-bioelectronic device translates the events of charged molecules into amplified electrical signals and outlines a generic format for the future exploitation of rich biological tunability and light-biomatter interplay for innovative bioanalytics and beyond.

Front Cover: Isothermal Rolling Circle Amplification and Lanthanide‐Based FRET for Femtomolar Quantification of MicroRNA (Anal. Sens. 6/2022)

The front cover picture illustrates rolling circle (RCA) amplification of one microRNA into a long DNA concatemer and its multicolor detection via terbium (Tb) and europium (Eu) Förster resonance energy transfer (FRET). This sensing concept was used to develop wash-free diagnostic assays on two commercial fluorescence plate readers. More information can be found in the Research Article by N. Hildebrandt and co-workers.

Aptamer-mediated DNA concatemer functionalized magnetic nanoparticles for reversible capture and release of circulating tumor cells

Effectively capturing, releasing, and reanalyzing circulating tumor cells (CTCs) are critical in cancer diagnosis and individualized treatment. Traditional immunomagnetic separation has disadvantages of low sensitivity and specificity, and is time-consuming and costly in CTCs capture. It is also easily disturbed by the microenvironment in releasing and analyzing CTCs. Here, we proposed an aptamer-mediated DNA concatemer functionalized magnetic nanoparticles (MNPs-AMDC) for the reversible capture and release of CTCs. In this study, aptamers were used both for efficiently capturing CTCs without complicated assembly steps and stimulus-response switch for releasing CTCs with little influence on cellular activity. The MNPs-AMDC was demonstrated to effectively capture (83%) and release CTCs with a good viability rate (92%). Moreover, this device was also tested in clinical blood samples, which would provide a universal tool for diagnosing cancer and treating individuals.

A Multiple-Target Simultaneous Detection Method for Immunosorbent Assay and Immunospot Assay.

Enzyme-linked immunosorbent assay (ELISA) is one of the most common methods in biological studies, and enzyme-linked immunospot (ELISpot) is a method to measure specific cell numbers by detecting protein secretion at a single-cell level. However, these two current methods can only detect one signal at one time and the sensitivity is not high enough to test low-concentration samples, which are major shortcomings in systematically analyzing the samples of interest. Herein, we demonstrated fluorescence-based oligo-linked immunosorbent assay (FOLISA) and fluorescence-based oligo-linked immunospot (FOLISPOT), which utilized DNA-barcoded antibodies to provide a highly multiplexed method with signal amplification. Signal amplification and simultaneous multiple-target detection were achieved by DNA complementary pairing and modular orthogonal DNA concatemers. By comparing FOLISA with traditional ELISA and comparing FOLISPOT with traditional ELISPOT, we found that the detection sensitivities of FOLISA and FOLISPOT are much higher than those of traditional ELISA and ELISPOT. The detection limit of ELISA is around 3 pg/mL, and the detection limit of FOLISA is below 0.06 pg/mL. FOLISPOT can detect more spots than ELISPOT and can detect targets that are undetectable for ELISPOT. Furthermore, FOLISA and FOLISPOT allowed sequential detection of multiple targets by using a single dye or multiple dyes in one round and sequential detection in multiple rounds. Thus, FOLISA and FOLISPOT enabled simultaneous detection of a large number of targets, significantly improved the detection sensitivity, and overcame the shortcomings of ELISA and ELISPOT. Overall, FOLISA and FOLISPOT presented effective and general platforms for rapid and multiplexed detection of antigens or antibodies with high sensitivity, either in laboratory tests or potentially in clinic tests.

Highly sensitive electrochemical aptasensor for SARS-CoV-2 antigen detection based on aptamer-binding induced multiple hairpin assembly signal amplification

In this work, a brief electrochemical aptasensor was developed for highly sensitive detection of SARS-CoV-2 antigen utilizing an aptamer-binding induced multiple hairpin assembly strategy for signal amplification. In the presence of SARS-CoV-2, a pair of aptamers was brought in a close proximity according to the aptamer-protein antigen binding, which initiated strand displacement reaction thereby triggering a multiple hairpin assembly to obtain long linear DNA concatemers on the electrode surface. As the fabricated hairpin probes were labeled with biotin, massive streptavidin-alkaline phosphatases (ST-ALP) could be further introduced on the electrode interface via biotin-streptavidin interaction thus generating strong electrochemical signal in electrolyte solution containing 1-naphthol phosphate. Benefiting from the non-enzymatic multiple hairpin assembly signal amplification strategy, the designed aptasensor for SARS-CoV-2 spike protein detection exhibited the wide linear range from 50 fg·mL−1 to 50 ng·mL−1 and low detection limit of 9.79 fg·mL−1. Meaningfully, this proposed electrochemical assay provided a potential application for the point of care analysis of viral diseases under ambient temperature.

The Beauty of Bacteriophage T4 Research: Lindsay W. Black and the T4 Head Assembly.

Viruses are biochemically complex structures and mainly consist of folded proteins that contain nucleic acids. Bacteriophage T4 is one of most prominent examples, having a tail structure that contracts during the infection process. Intracellular phage multiplication leads to separate self-directed assembly reactions of proheads, tails and tail fibers. The proheads are packaged with concatemeric DNA produced by tandem replication reactions of the parental DNA molecule. Once DNA packaging is completed, the head is joined with the tail and six long fibers are attached. The mature particles are then released from the cell via lysis, another tightly regulated process. These processes have been studied in molecular detail leading to a fascinating view of the protein-folding dynamics that direct the structural interplay of assembled complexes. Lindsay W. Black dedicated his career to identifying and defining the molecular events required to form the T4 virion. He leaves us with rich insights into the astonishingly precise molecular clockwork that co-ordinates all of the players in T4 assembly, both viral and cellular. Here, we summarize Lindsay’s key research contributions that are certain to stimulate our future science for many years to come.

Reliable quantification of Cytomegalovirus DNAemia in Letermovir treated patients

Polymerase chain reaction (PCR) based methods are a fast and sensitive approach to detect and monitor viral load in Cytomegalovirus (CMV) patients. Letermovir (LMV) acts at a late stage during the CMV replication cycle and does not inhibit CMV DNA replication per se. Therefore, quantitative nucleic acid amplification testing might lead to the overestimation of viral load in patients treated with LMV and underestimate treatment success. To study this discrepancy, we treated infected cells with LMV or Ganciclovir (GCV) and compared viral progeny DNA levels. Prior to nucleic acid extraction and qPCR measurements we pretreated cell lysates and cell culture supernatants from infected cells with DNase I. This step assumes the degradation of DNA which is not protected from a viral capsid. LMV treatment did not reduce genomic copies (GC) in samples from whole cell lysates compared to samples treated with GCV. DNase treatment prior to DNA extraction, decreased GC in the LMV treated group to comparable levels as seen in the GCV group. In cell culture supernatants, LMV or GCV treatment led to an equivalent reduction of CMV GC. In this case, DNase treatment exerted a negligible effect on both groups. We conclude that the accumulation of concatemeric DNA within cells seems to be a confounding variable when monitoring LMV efficacy via qPCR. However, qPCR shows to be a reliable method to evaluate antiviral efficacy of LMV in cell free specimens. These results have strong clinical implications for the monitoring of CMV therapy during LMV treatment.

A novel electrochemical immunosensor for dibutyl phthalate based on Au@Pt/PEI-rGO and DNA hybridization chain reaction signal amplification strategy

Herein, a sensitive immunosensor for the rapid detection of Dibutyl phthalate (DBP) small molecules was developed. Its principle is based on Au@Pt/Polyethyleneimine (PEI)-rGO and hybridization chain reaction signal amplification system. First, Au@Pt/PEI-rGO modified electrode is used as a signal amplification platform and the antibody is fixed. In the presence of target DBP, it competes with DBP-bovine serum albumin-S0 for binding antibody. Finally, the DNA concatemer carrying a large number of signal molecules methylene blue combines with its S0 to produce a reduced electrochemical signal. Under the most reasonable conditions, this immunosensor showed a good linear range from1 pg mL−1 to 0.1 μg mL−1with a low detection limit of 0.276 pg mL−1. It has been verified that this immunosensor has good selectivity, reproducibility and stability. The designed biosensing solution will open up a new path for DBP monitoring and other small molecule targets in the food field.

Complete Genome Sequence of Herpes Simplex Virus 2 Strain G.

Herpes simplex virus type 2 (HSV-2) is a common causative agent of genital tract infections. Moreover, HSV-2 and HIV infection can mutually increase the risk of acquiring another virus infection. Due to the high GC content and highly repetitive regions in HSV-2 genomes, only the genomes of four strains have been completely sequenced (HG52, 333, SD90e, and MS). Strain G is commonly used for HSV-2 research, but only a partial genome sequence has been assembled with Illumina sequencing reads. In the current study, we de novo assembled and annotated the complete genome of strain G using PacBio long sequencing reads, which can span the repetitive regions, analyzed the ‘α’ sequence, which plays key roles in HSV-2 genome circulation, replication, cleavage, and packaging of progeny viral DNA, identified the packaging signals homologous to HSV-1 within the ‘α’ sequence, and determined both termini of the linear genome and cleavage site for the process of concatemeric HSV-2 DNA produced via rolling-circle replication. In addition, using Oxford Nanopore Technology sequencing reads, we visualized four HSV-2 genome isomers at the nucleotide level for the first time. Furthermore, the coding sequences of HSV-2 strain G have been compared with those of HG52, 333, and MS. Moreover, phylogenetic analysis of strain G and other diverse HSV-2 strains has been conducted to determine their evolutionary relationship. The results will aid clinical research and treatment development of HSV-2.

Unified-amplifier based primer exchange reaction (UniAmPER) enabled detection of SARS-CoV-2 from clinical samples

Primer exchange reaction (PER) is an emergent method for non-templated synthesis of single stranded DNA molecules. PER has been shown to be effective in cell imaging systems and for detection of macromolecules. A particular application of PER is to detect a specific target nucleic acid. To this endeavor, two coupled DNA hairpins, a detector and an amplifier, play in accordance to extend a target nucleic acid with a concatemer DNA sequence. Here we introduced unified-amplifier based primer exchange reaction (UniAmPER) that beneficially extends the target by a unified-amplifier. The unified-amplifier operates as both detector and amplifier hairpins. The extension resulted in synthesis of concatemer G-rich sequences. The G-rich sequences were expected to form G-quadruplex (GQ) structures. Presence of the GQ structures were investigated by peroxidase activity of GQs in presence of hemin, H2°2 and 3,3′,5,5′-Tetramethylbenzidine (TMB) as well as by fluorescence signal generation upon intercalation of thioflavin T (ThT). The presented unified-amplifier in this study facilitates application of PER systems for development of colorimetric or fluorogenic biosensors. As a proof of principle, the method has been applied for detection of reversely transcribed cDNAs from clinical SARS-CoV-2 samples.

Preparation of long single-strand DNA concatemers for high-level fluorescence in situ hybridization

Fluorescence in situ hybridization (FISH) is a powerful tool to visualize transcripts in fixed cells and tissues. Despite the recent advances in FISH detection methods, it remains challenging to achieve high-level FISH imaging with a simple workflow. Here, we introduce a method to prepare long single-strand DNA concatemers (lssDNAc) through a controllable rolling-circle amplification (CRCA). Prepared lssDNAcs are used to develop AmpFISH workflows. In addition, we present its applications in different scenarios. AmpFISH shows the following advantages: 1) enhanced FISH signal-to-noise ratio (SNR) up to 160-fold compared with single-molecule FISH; 2) simultaneous detection of FISH signals and fluorescent proteins or immunofluorescence (IF) in tissues; 3) simple workflows; and 4) cost-efficiency. In brief, AmpFISH provides convenient and versatile tools for sensitive RNA/DNA detection and to gain useful information on cellular molecules using simple workflows.

Highly Selective and Sensitive microRNA-210 Assay Based on Dual-Signaling Electrochemical and Photocurrent-Polarity-Switching Strategies.

Highly sensitive and selective microRNA (miRNA) assay is of great significance for disease diagnosis and therapy. Herein, a magnetic-assisted electrochemistry (EC)-photoelectrochemistry (PEC) dual-mode biosensing platform was developed for miRNA-210 detection based on dual-signaling EC and photocurrent-polarity-switching PEC strategies. Porous magnetic Fe3O4 octahedra with a large surface area were synthesized by calcining Fe-based metal-organic frameworks. Subsequently, the magnetic photoelectric materials (Fe3O4@CdS) were developed by the successive ionic layer adsorption and reaction method in Cd2+ and S2- solutions. Then, the self-assembled DNA nanoprisms contained three thiols/hanging arms that could capture miRNA-210 efficiently and were anchored to the Fe3O4@CdS octahedra via the Cd-S bond. When miRNA-210 was present, the double-stranded DNA concatemers [the self-assembled duplex helixes based on a pair of methylene blue (MB)-labeled single-stranded DNAs (AP1 and AP2) through the hybridization chain reaction and then intercalated with adriamycin (Dox) into their grooves] were connected with the Fe3O4@CdS-DNA nanoprisms. MB and Dox not only acted as the electrochemical probes but also synergistically switched the photocurrent polarity of the Fe3O4@CdS octahedra. Thus, miRNA-210 was assayed sensitively and selectively via the proposed EC-PEC dual-mode biosensing platform. Additionally, the abovementioned recognition steps occurred in a homogeneous system, and the effects of the impurities and interferences on the miRNA-210 assay could be easily avoided by magnetic separation due to the good magnetic properties of Fe3O4 octahedra. The proposed EC-PEC dual-mode biosensing platform showed a wide range of potential applications in bioanalysis and early diagnosis of disease.