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Try out PMC Labs and tell us what you think. Learn More. Recently, aptamers have attracted attention in the biosensing field as signal recognition elements because of their high binding affinity toward specific targets such as proteins, cells, small molecules, and even metal ions, antibodies for which are difficult to obtain. Aptamers are single oligonucleotides generated by in vitro selection mechanisms via the systematic evolution of ligand exponential enrichment SELEX process. In addition to their high binding affinity, aptamers can be easily functionalized and engineered, providing several signaling modes such as colorimetric, fluorometric, and electrochemical, in what are known as aptasensors. In this review, recent advances in aptasensors as powerful biosensor probes that could be used in different fields, including environmental monitoring, clinical diagnosis, and drug monitoring, are described. Advances in aptamer-based colorimetric, fluorometric, and electrochemical aptasensing with their advantages and disadvantages are summarized and critically discussed. Additionally, future prospects are pointed out to facilitate the development of aptasensor technology for different targets. In this context, the development of fast, simple, low-cost, high-sensitivity, and specific sensors for detecting pollutants or early stages of diseases is important. Because of the great advances in molecular biology and genetic engineering, the use of RNA and DNA has expanded not only in biology, for storing and transmitting genetic information, but also in the identification of antibiotics, proteins, peptides, amino acids, and even small molecules. Gold et al. Aptamers consist of 3D-folded structures of single-stranded oligonucleotides with lengths of usually 20—60 bases of nucleotides selected in vitro via the systematic evolution of ligand exponential enrichment SELEX process. The SELEX process is applicable for single targets, complex target structures, or even mixtures without proper knowledge of their composition. SELEX can be used to select aptamers with high affinities and specificities for their targets and low dissociation-constant values, across the low nanomolar to picomolar range. Aptamers can be selected through the in vitro process independent of animals or cell lines. The selection of aptamers for toxic target molecules or molecules with no or low immunogenicity is possible. Different modifications can be introduced in the basic SELEX process for the selection of the desired aptamer specifications for a specific application. The aptamers can be functional for native conformations of target molecules on live cells, so cell surface transmembrane proteins can be considered as targets \\\\\\\\\\\\\\\[ 3 , 4 , 5 , 6 \\\\\\\\\\\\\\\]. The 3D folded structure permits the formation of stable complexes with various targets, such as proteins, nucleic acids, and small molecules \\\\\\\\\\\\\\\[ 7 , 8 , 9 , 10 , 11 , 12 , 13 \\\\\\\\\\\\\\\]. Compared to antibodies, these aptamers are characterized by numerous advantages such as a wide range of targeted molecules inorganics, organics, cells, viruses, and bacteria, among others , facile preparation on a massive scale, low molecular weights, high temperature and pH stability, easier modification, and long-term storage stability. These advantages promote their importance and application in many fields such as biosensing, therapy, and diagnostics \\\\\\\\\\\\\\\[ 14 , 15 , 16 , 17 , 18 , 19 , 20 , 21 , 22 , 23 \\\\\\\\\\\\\\\]. Because of their size-dependent physical and chemical properties, biocompatibility, electronic properties, magnetic properties, and ability to manifest biological signaling and transduction mechanisms, the nanomaterials are providing great advances in the field of sensors. Several nanomaterials have been synthesized with different sizes and shapes such as gold nanoparticles AuNPs , silica nanoparticles, silver nanoparticles AgNPs , magnetic nanoparticles MNPs , and carbon quantum dots CQDs as unique templates for many applications such as biomaterial assays; the diagnosis, monitoring, and treatment of disease; and drug delivery \\\\\\\\\\\\\\\[ 24 , 25 , 26 , 27 , 28 , 29 , 30 , 31 , 32 , 33 , 34 , 35 , 36 , 37 , 38 \\\\\\\\\\\\\\\]. Interestingly, nanomaterials are being used as potential candidates for immobilization using aptamers to obtain nanosensor probes for several targets, with more amplification and various signals such as colorimetric, fluorometric, electrochemical, or optical \\\\\\\\\\\\\\\[ 31 , 39 , 40 , 41 , 42 , 43 , 44 , 45 , 46 , 47 , 48 , 49 \\\\\\\\\\\\\\\]. Sensor fabrication mainly depends on target recognition and signal transduction. Thus, nanomaterials provide desirable signal transduction for converting molecular recognition events into physically detectable fluorescent, colorimetric, and electrochemical signals \\\\\\\\\\\\\\\[ 50 , 51 \\\\\\\\\\\\\\\]. Simultaneously, nanomaterials play a significant role regarding an increase in the immobilization density and orientation of aptamers due to the high surface areas of nanomaterials and, thus, provide high binding capacity for targets \\\\\\\\\\\\\\\[ 52 , 53 , 54 \\\\\\\\\\\\\\\]. However, the high accumulation leads to the restriction of the 3D-structure formation. Because of that, the aptamer density on the nanomaterial surface needs to be precisely optimized \\\\\\\\\\\\\\\[ 55 , 56 \\\\\\\\\\\\\\\]. This systematic review is focused on recent approaches to aptamer engineering for biorecognition for several objectives. Herein, previous and current advances related to aptamer-based sensing protocols are provided, highlighting the possible detected signals, with a focus on the use of different nanomaterials with distinct configurations. The most significant studies on aptasensor development have been collected, providing the possible strategies available for using aptasensors. Moreover, a deep discussion on colorimetric, fluorescence, and electrochemical detection strategies is provided. Among the available assay recognition signals, colorimetric methods are considered simple and efficient with a great potential for point-of-care diagnostics, as the detection responses are simply visually discerned by the naked eye using simple, low-cost, and effective instrumental techniques. The basic colorimetric strategies involve the detection of the target by a color change through the naked eye and simple instrumentation. Colorimetric biosensors based on aptamers have demonstrated their sensitivity and selectivity, in addition to their effective potential for rapid onsite diagnosis without complicated instrumentation. However, the colorimetric aptasensor has some limitations such as the influence of color from samples; the time-consuming nature of the fabrication process \\\\\\\\\\\\\\\[ 57 , 58 \\\\\\\\\\\\\\\]; the difficulty in employing it in multiple-target assays, which are highly demanded in clinical diagnosis \\\\\\\\\\\\\\\[ 59 \\\\\\\\\\\\\\\]; and the small range of optimized pH solutions \\\\\\\\\\\\\\\[ 60 \\\\\\\\\\\\\\\]. Nobel metal nanomaterials, in particular, AuNPs and AgNPs, are excellent signal transducers for colorimetric analysis due to their significant optical properties associated with their particle size, size distribution, and shape \\\\\\\\\\\\\\\[ 61 , 62 , 63 \\\\\\\\\\\\\\\]. AuNPs are used for color-change-based detection, in which the AuNPs are employed as nanoassembly units for immobilization with aptamers for constructing a colorimetric aptasensor because of their unique features, including simple synthesis and unique optical, thermal conductivity, and electronic properties \\\\\\\\\\\\\\\[ 65 , 66 , 67 , 68 \\\\\\\\\\\\\\\]. Furthermore, the large specific surface area facilitates numerous adsorptions of biomacromolecules onto AuNP surfaces via electrostatic interaction, protecting them against aggregation and rendering them a good signal transducer for aptasensor construction \\\\\\\\\\\\\\\[ 70 , 71 \\\\\\\\\\\\\\\]. Based on the unique colorimetric capabilities of the distance-dependent surface plasmon resonance of AuNPs, a great variety of label-free colorimetric bioassay strategies have been developed. The color of AuNPs is extremely sensitive to their dispersion and aggregation in a solution, comprising the interparticle plasmon coupling changing, inducing SPR shifts \\\\\\\\\\\\\\\[ 72 \\\\\\\\\\\\\\\]. However, there are some limitations in using a colorimetric sensor based on the SPR response other than the common disadvantages of colorimetric sensors previously mentioned, including the fact that the SPR sensors are prone to interference due to not only the absence of a response to a change in the refractive index, but also non-specific binding that can induce interference \\\\\\\\\\\\\\\[ 73 , 74 \\\\\\\\\\\\\\\], leading to false results and limiting their applicability. In some cases, variations in the plasmonic response depending on the location of the target on the surface of the nanoparticles \\\\\\\\\\\\\\\[ 75 , 76 \\\\\\\\\\\\\\\] are detected. Moreover, large nanoparticle aggregates are unstable in solution, which could lead to some measurement error \\\\\\\\\\\\\\\[ 77 , 78 , 79 \\\\\\\\\\\\\\\]. In this regard, the binding interaction induces a change in the refractive index around the surface of the AuNP that modulates its resonance angle. Therefore, the SPR has been used in an aptamer-based colorimetric strategy for the assay of several targets \\\\\\\\\\\\\\\[ 80 , 81 , 82 , 83 \\\\\\\\\\\\\\\]. Gupta et al. In this assay, the AuNPs were covered with graphene oxide GO , followed by functionalization with a specific aptamer for E. In the presence of the E. Moreover, Lai et al. In the presence of malathion, the specific aptamer departs from the AuNP surface to bind with the malathion target, leaving the AuNP surface unprotected and tending to aggregate in a highly concentrated NaCl solution, with a distinct color change from red to blue. For more signal amplification, Lai et al. Chen et al. Finally, the hybridization between the amplicons and the AuNP probe took place for 5 min before adding MgSO 4 to induce the aggregation. The system showed a linear relationship for S. Similarly, a colorimetric aptasensor based on the aggregation of AuNPs as a signal transducer using cationic perylene probe as gold aggregating agent CPP was also provided by Lerdsrias et al. B Schematic protocol of the dopamine assay via aptamer-assisted AuNP-induced NaCl salt aggregation with two possible mechanisms of sensing, reproduced from \\\\\\\\\\\\\\\[ 91 \\\\\\\\\\\\\\\]. D Schematic principle of S. A Schematic illustration of the colorimetric aptamer sensor for E. C Schematic procedure for an S. The question is what the role of these targets and others in the stability and instability of AuNPs in solution is, and if this could influence the salt-induced AuNP aggregation or not, affecting the reliability of the proposed strategies. Recently, Zong et al. Liu et al. To confirm the competition, a proposed competition between two different targets i. This study reveals that both the aptamer and dopamine have a strong affinity to the AuNP surface, and the dopamine itself induces different degrees of aggregation based on its concentration. A more interesting finding is that the dopamine adsorption on the AuNPs could inhibit the aptamer adsorption, revealing that the most probable mechanism is the one described in Figure 2 Bb. Finally, it was disclosed that the system based on AuNP aggregation-assisting aptamer is strongly kinetically controlled depending on the degree of the binding affinity and competition between the AuNP and both the target and aptamer and, also, depending on whether this target can also induce the stabilization or destabilization of AuNPs \\\\\\\\\\\\\\\[ 91 \\\\\\\\\\\\\\\]. Based on this study, the free target in solution could cause problems for the sensor reliability, as also proposed by Zong et al. The AuNPs were confined for using in a colorimetric aptasensor with different strategies other than aggregation, depending on the catalytic performance of the NPs and anisotropic growth of the AuNPs. In the presence of S. Wang et al. Furthermore, Seongjae Jo et al. The changes in the LSPR, in the presence of cortisol and thrombin, showed linear ranges of 0. A Schematic illustration of the cortisol assay based on localized surface plasmon resonance LSPR aptasensor, reproduced from \\\\\\\\\\\\\\\[ 99 \\\\\\\\\\\\\\\]. B Scheme representing the thrombin assay via sandwich colorimetric solid-phase aptasensor based on the enhanced LSPR, reproduced from \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\]. C Schematic diagram showing the procedure and mechanism of the ATP assay via colorimetric aptamer technology based on the peroxidase-like catalysis properties of Fe 3 O 4 , reproduced from \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\]. Magnetic nanoparticles MNPs have also been used as a nanoassembling template for bioassay targeting, either alone or in combination with other nanostructures, where they can be easily isolated from the solution via an external magnetic field \\\\\\\\\\\\\\\[ 39 , , , , \\\\\\\\\\\\\\\]. Miao et al. The Fe 3 O 4 nanoparticles incorporated with aptamers were used for assaying trace levels of adenosine triphosphate ATP biomarkers colorimetrically, as provided by Li et al. In the presence of an ATP target, the target binds with its aptamer, which departs from the surface of the Fe 3 O 4 , decreasing its activity, with a detection range of 0. Replacing the antibodies with aptamers in the ELISA protocol has attracted attention for assaying food-borne pathogens e. First, the biotin aptamer was immobilized on the surface of an avidin-coated microplate to capture the E. The detection aptamer was fabricated with Cu-MOF NPs through avidin—biotin affinity as a transducer signaling based on the peroxidase activity of the Cu-MOF NPs catalyzing the conversion of the colorless TMB into the oxidized blue structure and, then, yellow in the presence of the acid. Xing et al. Wei et al. C Schematic illustration outlining the multicolor and photothermal assay for prostate-specific antigen PSA via magnetic beads assisting aptamer separation, reproduced from \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\]. D Detection of E. Li et al. Coupling between the aptamer and hybridization chain reaction HCR amplification was considered based on distance visualized readout technology for quantitative assays Figure 5 D. The HCR amplified the signal fold after the E. The integration of both fluorescent materials fluorophore dyes and fluorescent nanoparticles such as upconversion nanoparticles UCNPs , GO, and CQDs and aptamers can produce high sensitivity and selectivity, and a rapid analysis strategy, making them useful candidates for fluorescence-aptasensor bioassays \\\\\\\\\\\\\\\[ , , \\\\\\\\\\\\\\\]. The recognition affinity between aptamers and analytes induces conformational changes in the aptamer. This process can trigger changes in the fluorescent emission properties of the fluorophore dye or fluorescent nanomaterials, owing to changes in the original environments of these materials. The design of fluorescent aptasensors requires the use of hairpin aptamers aptabeacons , which are labeled with either a fluorophore or a quencher. Forster resonance energy transfer FRET typically uses a donor fluorophore and an acceptor quencher material. These operations are based on the disparity in the fluorescence responses of the fluorophores as a function of the potential and unique aptamer—target binding and the conformational-change degree \\\\\\\\\\\\\\\[ , , \\\\\\\\\\\\\\\]. Some of the relevant fluorescent strategies were employed for the assaying of several targets based on integrated aptamers and fluorescent materials as described in the following Figure 6 , Figure 7 and Figure 8. Khan et al. C Schematic illustration of the zearalenone assay using a ratiometric fluorescent nanoprobe with dual emission at and nm, reproduced from \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\]. Schematic description of the A sensing platform for E. B Proposed fluorescence aptasensor for E. D Proposed scheme of ATP assay based on ratiometric fluorescence from the binding between the ATP and aptamer complexes, reproduced from \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\]. B Fluorometric assay for acetamiprid using exonuclease integrated with the aptamer protocol, reproduced from \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\]. D Dual-mode fluorescent aptasensor using both aptamer and a DNAzyme to assay ATP with two different mechanisms, fluorometric and colorimetric signals, reproduced from \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\]. E TTX assay based on the difference in fluorescence response of the berberine reporter, reproduced from \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\]. F Turn-on fluorescence aptasensor assay for chloramphenicol based on oligomer quencher release, reproduced from \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\]. Some studies have focused on detecting several targets based on the aptamer-competitive quenching of carbon quantum dots and graphene materials \\\\\\\\\\\\\\\[ , , , , , \\\\\\\\\\\\\\\]. Shirania et al. Tan et al. This system was applied for the assaying of zearalenone ZEN , which could quench green-emitting g-QDs linked to an aptamer via an electron transfer mechanism Figure 6 C , with a LOD of 7. Recently, Guo et al. In the presence of thrombin or ATP, the respective aptamer binds with its target, leading to the restoration of the fluorescence, with LODs of 1. A method for signal amplification was introduced by Wen et al. As outlined in Figure 6 G \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\], two aptamers were used to sandwich the protein, where the second aptamer was functionalized with azide, which, upon polymerization, produces an amplified fluorogenic signal LOD: 0. Fan et al. The hybridization of the aptamer with the target leads to the restoration of the fluorescence. Rare-earth-element-doped UCNPs can convert low-energy light into high-energy light with short wavelengths via the photon mechanism. UCNP materials are characterized by high quantum yields, narrow emission peaks, excellent photostability, long fluorescence lifetimes, environmental friendliness, low toxicity, bulky anti-Stokes shifts, and high resistance to photobleaching. Moreover, infrared light induces minimal photodamage to the biological targets without exciting the biological fluorophoric substance, which can be confined in the matrix \\\\\\\\\\\\\\\[ 80 , , , , \\\\\\\\\\\\\\\]. UCNPs were used in the development of fluorescence aptasensors for assaying different targets \\\\\\\\\\\\\\\[ , , , , \\\\\\\\\\\\\\\]. However, they still have some limitations such as the difficulty in distinguishing different target concentrations by the naked eye upon excitation by a laser beam at nm, where the target can be only be detected using instrumentation. This was achieved through the attachment of the aptamer to the WS 2 surface via van der Waals forces. In the presence of E. Therefore, part of the fluorescent intensity is restored as a function of E. Moreover, E. Basically, the dispersion and aggregation affinity were harnessed to produce a quencher for the fluorescent materials, as used in the assaying of chlorpyrifos CPF \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\]. Qiu et al. Once the sensor is exposed to the ATP, the binding between the ATP and aptamer complexes opens the hairpin structure and releases the anchor sequence, which hybridizes with the ssDNA and induces the turning off of the green fluorescence, while the red fluorescence is turned on, with a LOD of 0. As mentioned above, aptamers are sequences of nucleic acid, and because of that, they can be integrated with a DNA system for fluorescent signal amplification to improve the detection limit and, therefore, be used in the early diagnosis of diseases. These amplification processes include rolling-cycle amplification, strand displacement reactions, hybridization chain reactions, and hairpin DNA cascade hybridization reactions \\\\\\\\\\\\\\\[ , , , , , \\\\\\\\\\\\\\\]. Ning et al. The amplification of the signal using exonuclease I Exo I was also integrated with the fluorescent aptasensor for assaying AFB1 \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\]. Employing the same technology, the exonuclease integrated with an aptamer was used to assay acetamiprid \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\]. In the absence of acetamiprid, the specific aptamer hybridizes with the complementary DNA labeled with ferrocene cDNA—Fc , without a change in the fluorescence intensity after the addition of the exonuclease RecJf , while in presence of acetamiprid, it will bind with the aptamer, which leaves the cDNA—Fc, and then digested by the RecJf exonuclease, liberating the Fc to interact with cyclodextrin and initiating photoinduced electron transfer, as outlined in Figure 8 B \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\]. The positively charged TPE-TA binds one aptamer and aggregates, resulting in an amplified fluorescence. When the target exosomes are introduced, the aptamer preferentially binds with its target. The aptamers themselves can be modified with a fluorophore and quencher materials at both ends without affecting their binding affinity toward the targets, so a FRET aptasensor strategy can be easily constructed in the presence of targets inducing fluorescence quenching \\\\\\\\\\\\\\\[ , , \\\\\\\\\\\\\\\]. Kang et al. The presence of ATP induces attraction between the two strands of the aptamer, which induces the quenching of the fluorescence dye on one aptamer end via the quencher labeled on the other end of the aptamer. Lan et al. Zhang et al. Sharma et al. An electrochemical aptasensor was fabricated using an aptamer as a bioreceptor and an electrochemical transducer, which translated the target—aptamer affinity into a measurable electrochemical signal through potentiometry, voltammetry, amperometry, impedimetry, or electrochemiluminescence. The potentiometric approach involves the measurement of the potential between the probe and the reference electrode without any net charge transfer. The amperometric approach is based on applying a potential and allowing a redox reaction to occur. The signal is defined as the current between the electrode and the counter-electrode. The voltammetric strategy includes the sweeping potential over time and recording the corresponding current; this strategy is based on a three-electrode system, with working, reference, and counter electrodes. The impedimetric technique involves measuring the charge transfer rate on the surface of the electrode for a kinetic study. The electrochemical aptasensors were modified with various nanomaterials such as carbon-based nanomaterials, metal—organic frameworks MOFs , AuNPs, and polymers for signal amplification \\\\\\\\\\\\\\\[ , , , \\\\\\\\\\\\\\\]. The electrochemical aptasensor mainly depends on the interactions occurring on the surface of transducer as a result of the induced reaction between the target and its specific aptamer, providing amperometric or potentiometric electrochemical signals. Another technique is based on the increase in charge transfer resistance via the impedance technique \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\]. Because of the formation of the Au—S bond between the AuNPs and the modified thiol aptamer, competition on the surface of the GCE occurs between the amoxicillin and its aptamer, achieving a detection sensitivity of 0. A — D Schematic illustrations for the fabrication of electrochemical aptasensors. C A sandwich-type aptasensor for thrombin assay using CSPH hydrogel-modified electrode, reproduced from \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\]. A sandwich-type electrochemical aptasensor has been fabricated for a thrombin assay, as demonstrated in Figure 9 C. The sensor probe was fabricated on a conductive supramolecular polymer hydrogel CSPH modified electrode, on which the thrombin-binding aptamer 1 TBA1 was immobilized via amide bonds, while the thrombin-binding aptamer 2 was modified using magnetic nanoparticles MNP-TBA2 as signal amplification probes; the sandwich-type electrochemical aptasensor showed a linear range of 1 pM to 10 nM, with a LOD of 0. Recently, enzymes such as HRP, glucose oxidase, and alkaline phosphatase, and electroactive compounds such as QDs, ferrocene Fc , ferrocyanide, methylene blue MB , and Cd nanoparticles were successfully incorporated in electrochemical aptasensor technologies to be used as signal enhancers \\\\\\\\\\\\\\\[ , \\\\\\\\\\\\\\\]. A homogeneous electrochemical aptasensor was fabricated based on the departure of a target-responsive label, the electroactive dye methylene blue MB , from an aptamer-gated zeolitic imidazolate framework-8 ZIF-8 as a function of the thrombin concentration Figure 10 C , with a linear range of 1 fM to 1 nM and LOD of 0. Enzymatic catalysis was incorporated in an electrochemical aptasensor for signal amplification \\\\\\\\\\\\\\\[ , \\\\\\\\\\\\\\\]. Fu et al. Magnetic beads were fabricated with an H5N1 aptamer to enhance the selectivity, followed by immobilization with concanavalin A ConA , GOx, and AuNPs for signal enhancement in the glucose-fluid-inducing formation of gluconic acid. A — C Schematic protocols for electrochemical aptasensor based on the target-responsive label electroactive dye amplification strategy. C Thrombin assay based on target-responsive methylene blue MB release from the ZIF-8 surface as a function of the thrombin concentration, reproduced from \\\\\\\\\\\\\\\[ 13 \\\\\\\\\\\\\\\]. D Schematic protocols for an electrochemical aptasensor based on an enzymatic catalytic reaction, reproduced from \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\]. The electrochemical method was integrated with amplification strategies for assaying numerous targets, such as sulfadimethoxine, malathion, OTA, kanamycin, ampicillin, atrazine ATZ , and acetamiprid \\\\\\\\\\\\\\\[ , , , , , , , , \\\\\\\\\\\\\\\]. Sulfadimethoxine was assessed electrochemically with the assistance of nuclease amplification Figure 11 A , in which a gold surface of an electrode was immobilized with a DNA probe, and then hybridized with a specific aptamer for sulfadimethoxine, forming a dsDNA that inhibited their digestion via nuclease P1. The sulfadimethoxine aptamer leaves the DNA probe in the dsDNA in the presence of sulfadimethoxine, allowing the digestion of the dsDNA via nuclease P1 and inducing the amplification of the electrochemical signal, with a response range of 0. Malathion was detected with the assistance of exonuclease I Exo I as an electrochemical signal enhancer, which enhanced the signal by over two times, as shown in Figure 11 B. Moreover, the electrodeposition synthesis of PDA—AuNPs provided satisfactory biocompatibility and electrical conductivity for the sensor. Exo I was used to promote the autocatalytic cycling of malathion by enhancing the current change, with a linear response range of 0. OTA and ATZ were detected electrochemically and photoelectrochemically with the assistance of cycling amplification, with a linear range of 0. A — C Schematic protocols for electrochemical aptasensor integrated with signal amplification based on nuclease strategy. A Assaying of sulfadimethoxine using gold electrode fabricated with dsDNA consisting of DNA probe and specific aptamer for sulfadimethoxine inhibiting nuclease digestion, reproduced from \\\\\\\\\\\\\\\[ \\\\\\\\\\\\\\\]. The distinctive and impressive advantages of aptamers compared with antibodies permit them to be preferred in molecular diagnostics for a wide range of biomarkers. Aptamers have intrinsic advantages, such as their availability for both chemical modifications and conjugation with different labels, facilitating their ability to be used to construct a sensitive and highly selective platform sensor. In this review, recent advances in the different methods of employing aptasensors including colorimetric, fluorometric, and electrochemical strategies were discussed. The simplicity and small sizes of the aptamers combined with the versatile optical properties and large surface areas of nanomaterials leads to a platform with great potential for highly sensitive and selective biological recognition and signal transduction for various analytes such as metals, small molecules, toxins, proteins, cells, and bacteria with lower LODs and high sensitivity and selectivity. In this review, each detection method with its own different strategies was emphasized briefly with schematic designs and all the aptamer information, including the detection ranges of the discussed aptasensors, summarized in Table 1. Each aptasensor method has advantages, but the limitations of each method have to be considered before assaying the biomarkers. The optical strategies are considered promising assay methods, as a sensitive response could be achieved; however, some limitations should be considered before assigning a suitable strategy. The colorimetric aptasensor sensors are considered more promising for point of case testing POCT owing to naked-eye readout; however, their sensitivity fails to meet the required criteria owing to the higher LODs compared to other methods. Moreover, some limitations were demonstrated in the AuNP—salt-induced aggregation, in which the surface of the AuNP was easily accessible by several targets and aptamers, affecting the reliability of the sensor. This hurdle was overcome by using a strong capping agent \\\\\\\\\\\\\\\[ 93 , 94 , 96 \\\\\\\\\\\\\\\]. Despite the fact that the fluorescent aptasensor achieved a high sensitivity for the assay of different targets compared to colorimetric, it requires laboratories and clinical centers with infrastructure for achieving POCT diagnosis, and the limitations of the photobleaching of the fluorescent molecules over time, compromising their stability, is considered an obstacle regarding fluorescent aptasensor development. Generally, electrochemical aptasensors are rapid, easy, and higher in sensitivity compared to optical sensors, making them the best candidates for the on-site rapid assay of biomarkers. Biomarker detection based on aptamer functionalization still needs to overcome these limitations in order to be available for the multi-detection of metal ions, DNA, and proteins. Furthermore, the development of more specific aptamers is still needed, as is also integration into sensor platforms. Aptasensor platforms need more attention regarding 1 simultaneous multiple marker detection; 2 the long-term stability of biosensor assays; 3 direct assays in real sample matrixes; 4 understanding the nature of the binding competition between an aptamer and target on the surface of a nanomaterial, which could affect the sensor reliability; 5 more focus on the future development of in vivo aptamer sensing technology, the possible problems, and their solutions; and 6 more intensive research regarding the improvement of the POCT of biomarkers using aptasensors. The aforementioned hurdles need to be overcome quickly for the achievement of a reliable and selective detection of markers with ultrasensitivity, with affordable and portable on-site analytical devices. We are sure that the scientific community has the talent to offer solutions to these hurdles in order to design an affordable, easy-to-use nanomaterial-based electro-optical aptasensor integrated with rolling-cycle amplification technology. National Center for Biotechnology Information , U. Journal List Sensors Basel v. Sensors Basel. Published online Feb 2. Samy M. Author information Article notes Copyright and License information Disclaimer. Received Dec 22; Accepted Jan Abstract Recently, aptamers have attracted attention in the biosensing field as signal recognition elements because of their high binding affinity toward specific targets such as proteins, cells, small molecules, and even metal ions, antibodies for which are difficult to obtain. Keywords: aptamer, colorimetric aptasensor, fluorometric aptasensor, electrochemical aptasensor. Colorimetric Aptasensor Among the available assay recognition signals, colorimetric methods are considered simple and efficient with a great potential for point-of-care diagnostics, as the detection responses are simply visually discerned by the naked eye using simple, low-cost, and effective instrumental techniques. Open in a separate window. Figure 2. Table 1 Examples of application of aptasensors for quantitative detection. Colorimetric E. Figure 1. Figure 3. Figure 4. Figure 5. Figure 6. Figure 7. Figure 8. Electrochemical Aptasensor An electrochemical aptasensor was fabricated using an aptamer as a bioreceptor and an electrochemical transducer, which translated the target—aptamer affinity into a measurable electrochemical signal through potentiometry, voltammetry, amperometry, impedimetry, or electrochemiluminescence. Figure 9. Figure Conclusions and Future Outlook The distinctive and impressive advantages of aptamers compared with antibodies permit them to be preferred in molecular diagnostics for a wide range of biomarkers. Acknowledgments Samy M. Shaban acknowledges the support from Egyptian Petroleum Research Institute. Conflicts of Interest The authors declare no conflict of interest. References 1. Tuerk C. Ellington A. In vitro selection of RNA molecules that bind specific ligands. Cell Biol. Stoltenburg R. Zhuo Z. Ohuchi S. Open Access. Highly sensitive and specific detection of small molecules using advanced aptasensors based on split aptamers: A review. TrAC Trends Anal. A photoelectrochemical aptasensor for the determination of bisphenol A based on the Cu I modified graphitic carbon nitride. Ojha Y. Selection and characterization of structure-switching DNA aptamers for the salivary peptide histatin 3. Liu Z. 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