Self‐Propelled Nanoswimmers in Biomedical Sensing

Micro/nanomotors are nano‐devices that can convert energy into power to achieve self‐propulsion. Compared to ordinary nanoparticles, the driving force of nanomotors is their unique advantage. This driving force enables the controllable movement with defined direction and speed, allowing these nanoswimmers to perform more complex functions. Biomarkers are important parameters in clinical medicine and are closely related to disease prediction, diagnosis, and condition monitoring. Existing clinical biomarker monitoring and detection methods often require sophisticated equipment and complex operations. The use of micro/nanomotors is expected to greatly simplify these processes with better sensitivity and reliability. This article introduce the major classes of nanoswimmers based on their sources of energy and driving modes, as well as discuss how nanoswimmers can be used as nanosensors for biomarker detection.


Introduction
Nano-scale robots operating inside the human body to cure disease or repair wounds is a concept that has been frequently depicted in science fiction films. However, since the introduction of Paxton and his colleagues' nanorod design in 2004, which is capable of self-propulsion through the decomposition of hydrogen peroxide, [1] this once-fictional scene has slowly begun to materialize. Although it was not the origin of nanomotors, Paxton's nanomotor design is unique in that it enables autonomous movement in the surrounding environment without external driving forces, such as electric or magnetic fields, thus holding great significance of the nanomotor technology.
Hitherto, many different types of micro/nanomotors (MNMs) have been created, and each of them presents unique motion patterns. [2] Their self-propulsion mechanisms, such as DOI: 10.1002/adsr.202300056 catalytic chemical fuel decomposition that create gas bubbles (i.e., bubble propulsion), chemical concentration gradients (i.e., self-diffusiophoresis), and even motile biological cells (e.g., motile bacteria), have been studied. New MNMs are constantly being developed for advanced applications such as biosensing, drug delivery and environmental remediation.
As for biomedical applications, nanoparticles face numerous challenges due to the diverse and complex biological barriers present in the human body. For example, the tumor microenvironment poses several challenges, including the presence of abnormal vasculature and extracellular matrix, and high interstitial pressure. These barriers restrict the ability of nanoparticles to reach their target site, and significantly impacting their performance. [3] Owing to the autonomous driving force of MNMs, they are believed to have improved cell and tissue penetration, thus greatly increasing the efficiency of nanoparticles in contact with targets. This is extremely important in biosensing and targeted drug delivery.
Biomarkers hold great significance in detection and treatment of disease. In most cases, biomarkers can represent the state of disease in the human body directly. The earlier a disease is detected, the greater the possibility of a cure. This is important for cancer and cardiovascular disease, as early detection is of great significance for a successful treatment. [4] The early diagnosis of these diseases is mainly through the detection of biomarkers that is also required to monitor the progression and recurrence during the subsequent treatment. However, there are still various technical difficulties in the detection of biomarkers both early and reliable. For example, the concentration of biomarkers in early stages of tumor metastasis is extremely low and often masked with other proteins, which greatly increases the detection difficulty. [5] Therefore, it is very important to invent reliable and efficient biomarker detection technology. As a research hotspot in recent years, MNMs have unique characteristics: fast-moving speed, autonomous directional control, easy surface functionalization, and high payload. [6] MNMs can effectively separate and detect targets from complex mixtures and improve the accuracy of detection, thus offering great potential in biosensing.  [7a] Copyright 2012, American Chemical Society. B) Schematic diagram of the bubble acts as a power within the microtubule and the expanded area inside the microtubule. Reproduced with permission [7b] Copyright 2011, Royal Society of Chemistry. C) Schematic diagram of an Al-Ga/Ti nanomotor driven by hydrogen generation from water. The Al-Ga alloy is the dark hemisphere on the right and the Ti coating is the green hemisphere on the left. Reproduced with permission [9a] Copyright 2012, American Chemical Society. D) Schematic diagram of the Al/Pd spherical nanomotor with acid, alkali, and hydrogen peroxide solutions as fuels. The direction of motion using different fuels is also shown. Reproduced with permission [9b] Copyright 2012, Wiley-VCH. community started to turn its attention to nanomotors should accredit Richard Feynman's talk: "There's Plenty of Room at the Bottom" in 1959. His presentation stimulated scientists' interest in the research of the new generation of nanotechnology.
The purpose of designing nanomotors is to obtain nanoparticles that can overcome microfluidic dynamics at low Reynolds numbers. Nanoparticles that do not have extra power should obey the scallop theorem in a low Reynolds number environment. [2d] Therefore, the two most important elements for designing a nanomotor are: 1) Stable power, which helps nanomotor to break the constraints of Brownian motion; 2) Controllable direction, which helps nanomotor to reach the target. Only when the above two requirements are met can the nanomotor move according to the design requirements. The driving principle of the nanomotor directly determines these two elements. Accordingly, nanomotors can be broadly divided into two types: fuel-based and fuelfree nanomotors.

Bubble-Propulsion
There are two major methods to enable MNMs with bubbles propulsion. The first is that MNMs form bubbles on their surface through chemical reactions, such as platinum nanoparticles catalyzing H 2 O 2 to generate oxygen, which breaks the hydrogen bond network of water. The liquid-air interface formed on the surface of the nanomotor reduces the surface tension. Because the air bubbles are only generated at one end of the nanoparticle, an interfacial tension gradient along the length of the nanoparticle propels it forward. [1] The other method is to generate high-speed motion by providing reverse thrust to the nanoparticle through the air bubbles formed in a small space. [7] Different shapes and materials also have obvious effects on the speed and motion mechanism of the MNM. Common MNMs have the following shapes: truncated cone, [7a,8] cylinder, [7b] sphere, [9] and double truncated cone. [10] In one of the pioneering works, Wang and co-workers designed a tubular polyaniline (PANI)/Zn microrocket that generates hydrogen bubbles through the redox reaction from the inner zinc coating in an acidic environment ( Figure 1A). The micromotors obtained a strong driving force and achieved a speed over 100 body length/s. [7a] The speed of the motor depends on the acid concentration, and its lifetime is related to the dissolution rate of zinc. Therefore, the service life of the micromotor depends on the thickness of the zinc coating and the acid concentration of the environment. The lifespan is generally 10 s to 2 min. Mei and co-workers designed a cylindrical nanomotor that uses PPt to catalyze the decomposition of H 2 O 2 . [7b] H 2 O 2 is decomposed under the catalytic activity of Pt to generate oxygen bubbles ( Figure 1B). Then the oxygen bubbles aggregate to form micronsized bubbles and are discharged from the tube to provide driving force.

Figure 2.
A) The driving principle of Pt-Au nanorods. Reproduced with permission [14] Copyright 2006, American Chemical Society. B) (i) SEM image of polystyrene spheres coated with gold and nickel. (ii) Energy dispersive X-ray spectroscopic composition analysis of polystyrene spheres Green: Ni, red: Au. Reproduced with permission [15] Copyright 2010, American Chemical Society. C) (i) SEM image of Bare CaCO 3 . (ii) SEM image and schematic diagram of CaCO 3 Janus particle. (iii) Schematic diagram of the movement of CaCO 3 Janus particles toward HeLa cells in an acidic environment. Reproduced with permission [17] Copyright 2016, Springer Nature.
In another work, Wang and co-workers fabricated proteindetecting nanomotors using Ti/Ni/Au/Pt microtubes by catalyzing the decomposition of H 2 O 2 as the power. [11] Gold can also be used as a catalyst for H 2 O 2 . Minteer and co-workers used gold in nanotubes to catalyze the decomposition of H 2 O 2 as the power to propel nanomotors for DNA detection. [12] Another common shape for bubble-propelled nanomotors is Janus nanospheres. Wang and co-workers designed an Al-Ga/Ti binary alloy microsphere Janus nanomotor, [9a] which operates using only water as the chemical fuel ( Figure 1C). The catalytically active Al-Ga sphere is prepared through microcontact mixing of aluminum microparticles and liquid gallium. The exposed Al-Ga alloy hemisphere side ejects hydrogen bubbles upon contact with water, resulting in a strong peroxide directed propulsion thrust. In another work from Wang's group, an Al/Pd Janus micromotor was fabricated, which can take advantage of three different fuels to generate thrust. [9b] The micromotor was fabricated by wrapping half of an Al sphere with a Pd coating ( Figure 1D). The microsphere is propelled by the generation of hydrogen bubbles from the Al side in strongly acidic and alkaline environments, as well as by the production of oxygen bubble thrust at the Pd side in hydrogen peroxide media. Notably, the micromotor exhibits exceptional speed and longevity in diverse chemical environments and can adapt independently to the presence of new fuels without affecting its propulsion behavior. It can utilize three different fuel sources, including acid, alkali, and hydrogen, making it a highly versatile and efficient nanomotor for a wide range of potential applications.

Self-Phoresis
Self-driven MNMs are typically categorized as either self-electrophoretic or self-diffusiophoretic. [13] The selfelectrophoretic motor is comprised of two parts, a "source" region that contains chemical fuel to release ions, and a "sink" region that consumes ions. Such a distribution of ions generates an electric field that points from cations excess region to depleted region. MNMs move by electrophoresis when negatively charged colloidal particles exist. Mallouk and coworkers designed a self-electrophoretic catalytic nanorod. [14] The nanorods were fabricated by electroplating Pt and Au on the surface of anodic alumina nanowires (Figure 2A). When exposed to a solution containing H 2 O 2 , the oxidation reaction occurs at the Pt end (the anode), while the reduction reaction takes place at the Au end (the cathode). An electric field is formed around the nanorod due to the uneven distribution of protons. The nanorods will move along the direction of Pt end because of the electronegative metal surface. The speed of the nanorods can achieve up to 30.2 ± 4.0 μm s −1 . Posner and co-workers also employed a similar principle to fabricate spherical self-electrophoretic micromotors. [15] They coated half of the 5 μm diameter polystyrene with Au and the other half with Ni ( Figure 2B). Spherical micromotors made using this principle can achieve a moving speed of up to 15 μm s −1 . The Pt-Cu nanorods made by Sen and co-workers can selfelectrophoretically swim in Br 2 or I 2 solution and can be used to detect I 2 concentration in the solution. [16] A self-diffusiophoretic MNM typically consists of two parts, a "source" region that releases both cations and anions to maintain charge neutrality in the active area, and an "inert" region. The difference in diffusion rates between cations and anions generates an electric field that accelerates the slower diffusing ions and decelerates the faster diffusing ones, thus preserving the electroneutrality of the solution and propelling the MNMs. Schmidt and co-workers fabricated Janus particles by depositing a chromium (Cr) layer on one side of the CaCO 3 sphere ( Figure 2C). [17] In an acidic environment, the nanomotors are propelled by the release of Ca 2+ , HCO 3− , OH − and H + ions, which generate an electric field. The particles move along the acid gradient toward the target cell and gradually decrease in size due to the decomposition of CaCO 3 during the movement.  [18] Copyright 2014, Wiley-VCH. B)(i) Schematic diagram of the magnetic field driven helix nanomotor. The applied magnetic field is aligned with the permanent magnetic moment, and the rotating magnetic field controls the horizontal movement of the nanomotor. (ii) Schematic diagram of nanomotor control: The system was placed in a three-axis Helmholtz coil, and the motor movement in the cell was recorded and analyzed by light microscopy. Reproduced with permission [19] Copyright 2018, Wiley-VCH.

Fuel-Free Nanomotor
Although fuel powered nanomotors have the advantage of selfpropulsion without the need of external energy input, their maneuverability and potential toxicity of chemical fuels and the incompatibility of high concentrations of ions significantly limit their biomedical applications. For example, H 2 O 2 , a fuel commonly used in nanomotors, cannot exist in high enough concentrations in the human body to effectively drive many H 2 O 2decomposing nanomotors. Apart from chemical fuels, MNMs can also be driven by external energy or field, such as ultrasonic waves, [18] magnetic field, [19] light [20] and electrical field. [21] Compared with fuel-based MNMs, MNMs powered by external energy generally have better maneuverability and service life.
In an early work, Mallouk and co-workers designed gold nanorods whose motion was activated by ultrasound. [18] Gold nanorods can be internalized after co-incubation with HeLa cells (Figure 3A). The directional motion and rotation of the nanorods can be promptly controlled by adjusting the sound waves, highlighting its potential to controllably manipulate intracellular organelles. Then nanorods can be activated using 4 MHz resonant ultrasound, and the nanorods can move within the Hela cells without destroying them. Wang and co-workers developed a nanomotor for RNA detection, utilizing GO-AuNWs as the motor body propelled into cells by ultrasound stimulation. [22] Nevertheless, there are inherent limitations of ultrasonicdriven nanomotors, as they require sophisticated apparatus to generate ultrasound and require set conditions to form standing waves. These conditions may not always be applicable, especially in complex environments such as the human body.
Magnetic fields have shown certain advantages for powering nanoswimmers because of the ease of precise motion control and theoretically unlimited motion lifetime. Magnetically actuated nanomotors are often designed in a helical shape, as it enables them to move through a fluid environment more efficiently by creating a directional corkscrew-like motion. Wang and coworkers fabricated helical micromotors based on the bionic design of vascular plants. [23] After taking out the spiral xylem vessels in the plant, they plated a 20nm-thick titanium layer and an 80nm-thick nickel layer on the surface using an electron beam evaporator, and then cut them to the required length. This method mass-produced magnetic microswimmers with a diameter of 60 μm and a speed of up to 250 μm s −1 . Precise control of microswimmers is achieved using three orthogonal Helmholtz coils.
Ghosh and co-workers fabricated helical magnetic nanomotors using silica by the glancing angle deposition technique ( Figure 3Bi). [19] The nanomotor can be controlled by a lowamplitude (<80 Gauss) magnetic field rotating at a frequency of 1-5 Hz. Magnetic nanomotors can also be internalized by cells and can be suspended within cells under the control of a magnetic field (Figure 3Bii). Oksuz and co-workers designed dyelabeled single-stranded DNA (ssDNA probe) modified, magneticdriven Au-Ni nanomotors for intracellular miRNA detection with low detection limits of 2.9 and 1.6 pm for fluorescence and speed-based detection, as well as anti-cancer drug delivery. [24] Light-driven nanomotors have also become a popular research topic in biosensing. One benefit of employing light irradiation is the ability to utilize multiple properties of light, such as wavelength, polarization state, and orbital angular momentum, in (ii) Schematic diagram of micromotor movement by switching light and electrostatic interactions: particles aggregate under UV light due to electrostatic attraction between TiO 2 and Pt of adjacent motors; aggregated particles begin to disperse due to Brownian motion when UV is turned off; When the UV light is turned on again, the light-induced particle motion is accompanied by osmotic and electrophoretic repulsion, and the dispersed micromotors are further separated. Reproduced with permission [20a] Copyright 2016, Royal Society of Chemistry. B) (i) Schematic diagram of the movement of WO 3 /Ag MNM through self-electrophoresis/self-diffusion electrophoresis. (ii) Schematic diagram of the force of WO 3 /Ag MNM rotation. Reproduced with permission [26] Copyright 2022, Elsevier. C) Schematic diagram of the synthesis and working method of the Janus nanomotor driven by two methods. Reproduced with permission [27] Copyright 2022, American Chemical Society.
addition to the irradiation direction for controlling the movement of the nanomotors. This multichannel control provides a more comprehensive and precise way of navigating the nanomotors, there are two major strategies for designing light-driven nanomotors: 1) Fabrication of asymmetric nanostructures using photoactive materials; 2) Asymmetric exposure of photosensitive materials. [25] Janus structure is the most common choice for realizing structurally asymmetric light-driven nanomotors due to its structural anisotropy. The TiO 2 /Pt Janus nanomotor designed by Guan and co-workers uses light as the energy source and water as the fuel. [20a] Under the irradiation of UV light, the nanomotor will undergo a redox reaction with water. The Ti side undergoes an oxidation reaction and is positively charged, and the Pt side un-dergoes a reduction reaction and is negatively charged, thus resulting in opposite charges on the nanomotor (Figure 4Ai). The aggregation and dispersion of nanomotors can be controlled using continuous and pulsed UV irradiation ( Figure 4Aii). Compared with spontaneously reactive self-propelled nanomotors, light-driven nanomotors have longer lifetimes and better controllability.
Dong and co-workers synthesized Ag nanoparticle decorated WO 3 nanorods (WO 3 /Ag) using hydrothermal and surface modification methods, which can sense light changes and perform biomimetic motion. [26] WO 3 and Ag generated ionic and non-electrolyte O 2 gradients, respectively, due to the decomposition of H 2 O 2 in the photocatalytic solution ( Figure 4B). Based on the mechanism of self-electrophoresis/self-diffusion electrophoresis, this nanomotor can realize in situ rotational motion under vertical light and negative phototaxis when the direction of illumination changes.
A recent nanomotor design by Liu, Zhao and co-workers combines enzyme-based nanomotor with remote speed regulation capabilities by using near-infrared (NIR) light as "optical brakes" consisting of asymmetric domains, with a SiO 2 @Au core@shell nanosphere and a periodic mesoporous organosilica (PMO) domain co-modified with glucose oxidase (GOx) and catalase (CAT). [27] The GOx/CAT tandem pairs catalyze glucose decomposition to create a concentration gradient, producing a diffusiophoretic force toward the Au nanoshell side ( Figure 4C). Meanwhile, the Au nanoshell creates a thermal gradient under NIR light, driving the nanomotor via thermophoresis in the opposite direction, toward the PMO side. This design allows for remote regulation of nanomotor speed (3.46-6.49 μm s −1 , or 9.9-18.5 body-length/s) by adjusting the power density of the NIR light at different glucose concentrations, effectively acting as an "optical brake". This approach addresses the issue of uncontrollable nanomotor speed in stable physiological environments and has potential for various applications in nanotechnology.
An external electric field can also be used to control MNMs, and there is no fuel consumed during this process. Granick and co-workers used electric field to control the self-assembly of Janus Ti/Si O 2 spheres. [21a] When an electric field is applied, the silica particles and the half-metal-coated Janus particles interact due to dipoles. Therefore, the force between silica particles and Janus particles is greater than the repulsive force between Janus particles, resulting in chiral clusters with silica particles as the core (Figure 5a). Thus tetrahedrons (assembled with high activity) and square cones (assembled with low activity) are formed. Increasing the diameter of the central particle can also form pentamers and hexamers (Figure 5b). Further research by the team found that the self-assembly of particles can be affected by changes in the frequency, strength, and type of the electric field. [28]

Application of MNMs in Biomedical Sensing
The timely and accurate detection of specific biomarkers can realize the prevention, diagnosis, treatment, and monitoring of diseases. [29] Existing biosensor technology still faces several problems such as high detection cost, long detection time, and limited sensitivity. The introduction of MNMs as new biosensors is promising in addressing these issues ( Table 1).

Nucleic Acid
Nucleic acids, such as DNA and RNA, are biomolecules that store and transmit genetic information in living organisms. Due to their crucial role in maintaining cellular function, any changes in their structure or expression can be indicative of various biological processes and diseases, such as cancer, [58] infectious [59] and neurodegenerative [60] diseases. As such, nucleic acids can be used as biomarkers to detect and diagnose diseases, monitor treatment responses, and predict disease outcomes. In the presence of an AC electric field, one pure silica inert particle rises from the bottom and aggregates with three or four Janus active particles. At low activity and low electric field (40 V mm −1 ) aggregated to form tetrahedral rotors or high activity and high electric field (80 V mm −1 ) generated square cone rotors. The scale bar is 10 μm. B) The first row is 3 μm in diameter and the second (high activity, 80 V mm −1 ), third (low activity, 40 V mm −1 ) row is 6 μm in diameter. The scale bar is 3 μm. Reproduced with permission [21a] Copyright 2016, Wiley-VCH.

Oligonucleotide
Oligonucleotides are short chains of nucleotides of less than 20 base pairs and play a critical role in cellular function. Any changes in their levels or modifications can serve as biomarkers for various diseases and biological processes. Oligonucleotides can be measured in various biological samples, including blood, urine, and tissue samples, and can be used for diagnostic, prognostic, and therapeutic purposes.
The micromotors designed by Wang and co-workers can rapidly and selectively isolate target oligonucleotides from complex biological samples. [30] Ti/Ni/Au/Pt microtube rockets were decorated with binary self-assembled monolayers composed of a short-chain 6-mercapto-1-hexanol (MCH) and a short-chain 6mercapto-1-hexanol (MCH). The rapid movement of the microrockets in the presence of H 2 O 2 induces fluid convection, which  [30] Peroxide and sodium cholate Rapidly isolate samples directly from raw biological samples without preparation and washing steps.

DNA
Au-Pt nanomotor [31] H 2 O 2 Can be directly observed with an optical microscope.
Cu-Pt microtubule [12] H 2 O 2 Minimum detectable concentration needs to be improved.
Jellyfish-like Micromotor [33] H 2 O 2 Good motion performance, stability, and reproducibility in protein-rich biological media.
RNA ssDNA@GO-functionalized gold nanomotor [22] Ultrasound Fluorescent signals representing the concentration of target RNA can be easily observed with a microscope.
E. coli Au/Ni/PANI/Pt microengine [42] H 2 O 2 Captured E. coli can be visualized optically and differentiated from non-target cells.
Au-Ni-Au nanowire [43] Magnet field and ultrasound Chemical fuel free.
pH Magnetic Pebble [45] Magnet field Can be controlled without direct mechanical contact.
DNA micromotor [48] Urea Quantitative and dynamic pH sensing in a few microseconds.
Lipopolysaccharides Pt-F 3 O 4 Janus micromotors [55] H 2 O 2 or magnet field Can be directly observed under a microscope.

Iodine
Cu-Pt nanorod [16] Iodine The moving speed varies linearly with the iodine concentration.
Adv. Sensor Res. 2023, 2, 2300056  [31] Copyright 2010, Springer Nature. C) (i) DNA1 was immobilized on the microtubule surface by EDC-NHS and PtNP-DNA coupling. (ii) Detection principle of nanomotors. Reproduced with permission [12] Copyright 2015, Royal Society of Chemistry. D) (i) Schematic diagram of the assembly of the micromotor. (ii) Schematic diagram of the working principle of the micromotor. Reproduced with permission [32] Copyright 2017, Royal Society of Chemistry. E) Schematic diagram of the assembly and working principle of the jellyfish-like micromotor. Reproduced with permission [33] Copyright 2019, American Chemical Society.
enhances the DNA hybridization efficiency, thus enabling the rapid and selective isolation of nucleic acid targets ( Figure 6A). This protocol enables fast DNA isolation directly from raw biological samples without preparation and washing steps. The microrockets can also deliver target oligonucleotides to other regions for further analysis.

DNA
Variations or mutations in the DNA sequence, such as deletion, insertion, short tandem repeats (STRs), single nucleotide polymorphisms (SNPs), and other variations can lead to the development of diseases such as cancer, diabetes, and Alzheimer's disease. [61] However, there are several challenges in DNA detection: 1) The concentration of target DNA in the sample is too low; 2) Mapping is difficult due to the complexity of the human genome; 3) Rare sequences in DNA are difficult to be detected; and 4) Single nucleotides are too small for optical detection. [62] These problems above can be solved using nanomotors to enhance the separation and detection of target DNA in complex samples. The nanomotor designed by Wang and co-workers can detect the content of target DNA in the environment with a detection limit of 40 amol. [31] They linked dithiothreitol (DTT), thiolated capture probe (SH-CP) and mercaptohexanol (MCH) on the surface of the gold electrode. In the presence of target DNA, SH-CP-Ag nanoparticles (NPs) are captured due to complementary nucleic acid target binding (Figure 6Bi). H 2 O 2 was added after removing excess SH-DP-Ag NPs and the H 2 O 2 was decomposed due to the catalysis of Ag to provide power for the nanomotor (Figure 6Bii). The speed of the nanomotor is positively correlated with the number of Ag NPs attached to the gold electrode, which means the speed of the nanomotor is positively correlated with the amount of target DNA (Figure 6Biii). The movement speed of the nanomotor can be observed using an optical microscope to calculate the concentration of the target DNA.
Minteer and co-workers describe the design of a DNA sensor that uses tubular micromotors to detect the presence of DNA targets. [12] The sensor utilizes Pt nanoparticles conjugated to DNA as catalysts to drive the motion of the micromotors in a hydrogen peroxide solution. They conjugated DNA 1 to Pt NPs, and then attached the DNA 2 probe to the 11-mercaptundecanoic acid (MUA)-modified Poly(3,4ethylenedioxythiophene)-polystyrene sulfonate (PEDOT)-Au micromotors (Figure 6Ci). The presence of the DNA target recruits binding to Pt-DNA 1 nanoparticles, which are attached to the micromotor through DNA 2, causing the micromotor to move (Figure 6Cii). The motion intensity of the microtube sensor is directly related to the amount of the DNA target. This micromotor is not limited to DNA detection but can also be extended to the detection of other analytes, such as small molecules and proteins.
The strategies mentioned above are based on attaching metal NPs that can catalyze the decomposition of H 2 O 2 so nanoswimmers can increase their movement speed. Another idea for nanomotor biosensors is to design a speed reduction mechanism for the nanomotors by separating the catalysts attached to the nanomotors. In Wu and co-worker's study, multilayers of catalase are functionalized on the inside of a poly(3,4ethylenedioxythiophene and sodium 4-styrenesulfonate)/Au (PEDOT-PSS/Au) microtube using programmed DNA hybridization (Figure 6Di). [32] When exposed to target DNA, the sensing unit hybridized with it, leading to the release of the multi-layer DNA and multi-catalase and a decrease in motion speed (Figure 6Dii). By using the speed as a signal, the micromotor could detect DNA at concentrations ranging from 10 nm to 1 μm. Although the sensitivity is lower than the above two methods, the detection process is simpler, thus opening new avenues for designing simple and rapid biosensors for biomolecule detection.
Inspired by the jellyfish morphology, Ju and co-workers designed and synthesized umbrella-shaped micromotors. [33] A Janus multimetallic (Au/Ag/Ni/Au) shell was fabricated by sequentially sputtering layers of Au, Ni, Ag, and Au onto a SiO 2 microsphere ( Figure 6E). They then blocked the convex Au layer with 6-mercapto-1-hexanol (MCH) and dissolved the silica template. To create the sensing unit, they asymmetrically modified a sandwich DNA hybridization (DNA1/2/3) on the concave Au surface via Au-S binding. Further subsequent hybridization of complementary DNA strands on DNA3 was used to create long DNA complexes and covalently decorated catalase to form the power unit of the micromotor. Compared with nanotubes, surface modification of nano and micromotors using open space is easier and more efficient, and the identification of target molecules is also easier. The jellyfish-like micromotor exhibits good motion performance, stability, and reproducibility in motion-based DNA detection in protein-rich biological media. Therefore, it has considerable potential in biological sensing applications.

RNA
Having a key role in the regulation of different cellular processes, ribonucleic acid (RNA) is considered as an effective biomarker for disease diagnosis and prognosis. [63] MicroRNA(miRNA) is a single-stranded non-coding RNA molecule, and studies have shown that the abnormal expression of specific miRNAs is related to many diseases. [64] miRNA-21 is one of the earliest discovered mammalian miRNAs [65] and is also an important cancer biomarker. [66] In recent years, with the emergence of quantitative reverse transcription polymerase chain reaction (RT-qPCR), [67] RNA sequencing [68] and microarray, [69] the detection technology of RNA is relatively mature. But these techniques often require complex processes and expensive equipment, and the detection of RNA in body fluids often requires a larger volume of samples. Existing methods are also difficult to perform single-cell analysis and important information will lose when analyzing large numbers of cells simultaneously. [22] Wang and co-workers fabricated ultrasound-propelled singlestanded (ssDNA)@graphene oxide (GO)-functionalized gold nanomotors, which can achieve rapid and specific detection of mi-RNAs in single cells. [22] They used dye-ssDNA to modify the surface of GO-functionalized nanowires and GO quenched the fluorescence of the dye. Ultrasound was used to drive the nanowires into the cell, and the fluorescent dye was released from the surface of the nanowires when the target miRNA was hybridized with the dye-ssDNA (Figure 7A). This nanomotor enables rapid one-step intracellular biosensing of target miRNAs expressed in intact cancer cells at the single-cell level that has extremely low miRNA content.
In another study, Wang, Chen and co-workers developed an intracellular gene silencing strategy that employs acoustically propelled nanowires modified with interfering RNA (siRNA) payload ( Figure 7B). [34] The gold nanowires are wrapped with a Rolling Circle Amplification (RCA) DNA strand to anchor the siRNA, and the ultrasound-powered propulsion of the nanowires leads to fast internalization and rapid intracellular movement for an accelerated siRNA delivery and silencing response. The GFP/RCA wrapped AuNWs can be propelled rapidly into different cell lines and can dramatically accelerate the siRNA delivery and gene-messenger RNA (mRNA) silencing compared with static nanowires, achieving a 94% silencing response after a few minutes. The results suggest that DNA structures carried  [22] Copyright 2015, American Chemical Society. B) Schematic of the nanomotor-based gene silencing approach involves nanomotors that can move inside living cells, as seen in fluorescence and optical images. The schematic shows (i) the movement of GFP/RCA-AuNW nanomotors penetrating a HEK293-GFP cell under a US field and (ii) the natural process of gene-mRNA silencing inside living cells. Reproduced with permission [34] Copyright 2016, American Chemical Society. C) Schematic diagram of the detection steps. Reproduced with permission [24] Copyright 2021, American Chemical Society. by the US-propelled nanomotors for gene silencing represent an efficient tool that addresses the challenges associated with RNA transportation and intracellular delivery, offering potential as a co-delivery platform for therapeutics in gene therapy applications.
Oksuz and co-workers used magnetically driven gold-nickel nanowires combined with fluorescently labeled ssDNA to fabricate nanomotors that can detect intracellular miRNAs. [24] The nanomotor is driven into the cell by the magnetic force, and the ssDNA and miRNA begin to hybridize ( Figure 7C). During this process the fluorescence is quenched and the speed of the nanomotor is reduced. Results showed that the movement speed and the fluorescence intensity of the nanomotor are negatively correlated with the concentration of the target miRNA. The nanomotor can achieve a minimum detection of 1.6 pmol of miRNA-21.

Protein
Proteins are complex macromolecules that play crucial roles in various biological processes within the human body, including metabolism, immune responses, and cell signaling. As a result, changes in protein levels or function can often indicate the presence of diseases or disorders. As biomarkers, proteins have several advantages, including their high sensitivity and specificity, the ability to measure them in a variety of samples, and their potential for use in non-invasive diagnostic tests. Studies have shown that more than one thousand proteins have the potential to be biomarkers. [70] For example, C-reactive protein (CRP) for the detection of sepsis, -fetoprotein in hepatocellular carci-noma, and mucin-related glycoprotein in various cancers. [71] The enzyme-linked immunosorbent assay (ELISA) is a common and reliable method for detecting protein biomarkers. Taking advantage of the autonomous movement ability of nanomotors, recent studies have shown promising results in ELISA technology with more rapid and efficient protein biomarker detection.
Wang and co-workers used mercaptohexanol (MCH) and thiolated thrombin aptamer (SH-TBA) to modify the surface of goldcoated microtubes. [11a] Thrombin is captured by SH-tTBA when the H 2 O 2 -fueled micromotor moves within the sample enabling the separation of thrombin from complex samples ( Figure 8A). To achieve release after thrombin capture, they modified the nanomotors with a mixed-binding aptamer (MBA) containing ATP (ABA) and -thrombin (TBA)-binding aptamers. So that the nanomotors can release the thrombin in a solution in the presence of ATP. The ability of micromotors to separate targets from raw biological samples is greatly improved compared to existing technologies, while avoiding preparation and washing steps. Detection can be achieved within 20 min. In a further research, they designed microtubular micromotors based on molecularly imprinted polymers (MIPs) for protein capture and transport. [35] Bilayer of poly(3,4-ethylenedioxythiophene) (PEDOT)/Pt-Ni microengines and fluorescein isothiocyanate (FITC)-labeled avidin (Av-FITC) is used as a template to illustrate the concept ( Figure 8B). The avidin-imprinted polymeric layer selectively captured and concentrated the fluorescent-tagged protein onto the moving microengine, allowing extraction and isolation of Av-FITC from raw serum and saliva samples in 5-10 min. A maximum coverage of 90% of the sample surface can be achieved. In their concurrent work, self-propelled micromotors are fabricated for the capturing and transporting target proteins between the www.advancedsciencenews.com www.advsensorres.com  [35] Copyright 2013, Royal Society of Chemistry. C) Schematic representation of micromotors for protein immunoassays. Reproduced with permission [11b] Copyright 2013, Royal Society of Chemistry. D) Schematic diagram of the assembly and working principle of the micromotor for detecting carcinoembryonic antigen. Reproduced with permission [36] Copyright 2014, American Chemical Society. E). Schematic illustration of the magnetic micromotor immunoassay used to detect CRP. Reproduced with permission [37] Copyright 2020, Elsevier. different reservoirs of a lab-on-a-chip (LOC) device. [11b] This design can eliminate many time-consuming washing steps compared to ELISA bioassays, and the use of different microsphere tracers can also provide the possibility for multiplex immunoassays ( Figure 8C). Release of captured antigen after addition of elution solution also allows the micromotor to be recycled.
Ju and co-workers used a similar principle to design microtube motors that can directly detect target proteins. [36] Surfacemodified microtubes shuttle in the sample, and secondary antibody-modified glycidyl methacrylate microspheres (GMA) bind to the nanomotor in the presence of the target protein ( Figure 8D). The speed of the micromotor is thus slowed down. The speed changes of the micromotors can be observed under a microscope, and it can be employed as a sensor to detect carcinoembryonic antigens in the range of 1-1000 ng mL −1 within 5 min. C-reactive protein (CRP) is generally used as a biomarker for the diagnosis of sepsis. Existing CRP detection methods, such as fluorescence and chemiluminescence immunoassays or enzyme-linked immunosorbent assays require a large amount of sample reagents, and the process is cumbersome and timeconsuming. Using functionalized carbon-based microtube motors, Escarpa and co-workers showed rapid and accurate detection of C-reactive protein (Er = 1%) from minimal sample volumes (<10 μL) in an ELISA assay. [37] The three-layered micromotors utilize rGO for antibody sandwich functionalization (ELISA), Ni for magnetic guidance and stopped flow operations, and Pt NPs for catalytic bubble propulsion ( Figure 8E). The micromotor achieved a motion speed of 140 μm s −1 and detected C-reactive protein within 5 min.
Procalcitonin (PCT) is associated with the degree and severity of sepsis infection and can be used as a biomarker for the diagnosis and detection of sepsis, while PCT can also predict Figure 9. A) Schematic diagram of multifunctional Janus particle detection of PCT: (i) PCT is specifically captured in blood, (ii) signal generation mechanism: Adding H 2 O 2 creates air bubbles that propel the particles forward and disperse the color on the filter paper. Reproduced with permission [38] Copyright 2019, Elsevier. B) (i) Schematic diagram of the fabrication of the micromotor (ii) the process of PCT detection by fluorescence micromotorbased immunoassay (FMIm). Reproduced with permission [39] Copyright 2020, American Chemical Society. the risk of death in patients with severe infection and ventilatorassociated pneumonia. [72] Rica and co-workers used Janus particles to fabricate a colorimetric sensor for the detection of PCT in blood. [38] They mixed the antibody-modified particles in the sample and placed them on filter paper after incubation. Enzymatic substrate was added, and the particles were propelled forward by the captured catalase breaking down the substrate, and larger, fewer colored spots were observed. The relative pixel intensities of color blobs can be quantitatively calculated using a smartphone ( Figure 9A). The sensor can detect PCT in blood in 10-15 min, compared with 30 min for traditional methods. Escarpa and coworkers used microtubes to make biosensors for the detection of PCT. [39] Micromotors are propelled catalyzing H 2 O 2 to generate bubbles with magnetic guidance. A specific anti-PCT antibody is immobilized on the surface of the micromotor, and a detection antibody exists in the solution at the same time ( Figure 9B). After the two antibodies are connected by PCT, the fluorescence intensity is measured under the microscope after being collected using a magnet by taking advantage of the magnetic properties (nickel layer) of the nanomotor. This method allows the detection  [40] Copyright 2014, American Chemical Society. B) Schematic diagram of the construction of nanomotors and the detection of circulating tumor cells. Reproduced with permission [41] Copyright 2020, Elsevier.
of PCT over the entire clinically relevant range without the need for sample dilution.

Circulating Tumor Cells
Circulating tumor cells (CTCs) are tumor cells that have detached from the primary tumor and entered the bloodstream, which can then travel to other parts of the body and form secondary tumors, which often leads to rapid deterioration of the patient's condition. [73] As such, CTCs have emerged as a promising biomarker for the early detection and monitoring of tumor metastasis. [74] The presence of CTCs in the blood is significantly correlated with the prognosis and survival rate of various cancers, so it is extremely important to detect the number of CTCs in the blood of patients. However, CTCs are extremely rare in blood (about 1-100 CTCs per 10 9 blood cells), [75] therefore the enrichment of CTCs is necessary before detection.
Pang and co-workers used the layer-by-layer (LBL) assembly method to prepare fast-response immunomagnetic nanospheres (IMNs), which can efficiently and rapidly collect CTCs in blood ( Figure 10A). [40] The IMNs were modified with anti-epithelialcell-adhesion-molecule (EpCAM) antibodies, which were able to capture extremely rare tumor cells in whole blood with an efficiency of over 94% after only a 5 min incubation. The isolated cells remained viable and could be directly used for culture, reverse transcription-polymerase chain reaction (RT-PCR), and immunocytochemistry (ICC) identification. The authors demonstrated the successful application of IMNs in the isolation and detection of CTCs in cancer patients peripheral blood samples, even detecting a single CTC in the whole blood sample, indicating their potential as a promising tool for CTC enrichment and detection.
Mao and co-workers combined near-infrared actuation as well as fluorescence to magnetic nanomotors ( Figure 10B). [41] The nanomotor was fabricated by modifying ferro ferric oxide nanoparticles with a hierarchical silica shell and platinum nanoparticles, and then adding FITC and an EpCAM antibody for selective capture of CTCs. Under near-infrared (NIR) irradiation, the nanomotor can be driven by local thermal gradients caused by asymmetric distribution of Pt nanoparticles. The collection efficiency was enhanced by light-driven nanospheres moving within the blood, which could achieve a cell capture rate of up to 98.75%. Due to their fluorescent properties, they can be applied in the visual inspection of whole blood samples from clinical cancer patients.

Viruses and Bacteria
Viruses and bacteria can serve as biomarkers for various diseases, including cancers and infections. By detecting and analyzing the presence and behavior of these microorganisms, we can gain insights into the mechanisms of the diseases and develop new diagnostic and therapeutic strategies. Recent advances in MNMs have greatly improved the ability to identify and characterize viruses and bacteria as biomarkers, paving the way for more precise and personalized healthcare.
Wang and co-workers synthesized an Au/Ni/PANI/Pt microengine functionalized with the concanavalin A (ConA) lectin bioreceptor to isolate Escherichia coli (E. coli) from samples. [42] Lectin is bound to the gold outer surface of 8 μm long nanotubes through a self-assembled monolayer, and the microengine moves at a speed of 80um/s in the sample by decomposing H 2 O 2 . Lectins on the surface of nanotubes recognize E. coli cell walls through O-antigen structural binding. The capture of E. coli by the microengine can be clearly observed under a microscope Figure 11. A)(i) Schematic illustration of microengines for selectively picking up, transporting, and releasing E. coli. (ii) Schematic illustration of surface modification of microengines by lectin receptors. Reproduced with permission [42] Copyright 2012, American Chemical Society. B) Au-Ni-Au nanowire motors for selective capture and transport of biological targets. Reproduced with permission [43] Copyright 2013, American Chemical Society. C) Schematic diagram of nanomotor-based bead-motion cellphone (NBC) system for Zika virus detection. Reproduced with permission [44] Copyright 2018, American Chemical Society.
( Figure 11A). Compared with traditional microscopy, ELISA, PCR and other time-consuming and multi-step techniques, this method can quickly and efficiently isolate and detect E. coli from samples. In their subsequent work, nano-microtubes were improved to synthesize magnetically guided ultrasonic-driven Au-Ni-Au nanowire motors. [43] The motor is propelled acoustically by mechanical waves generated by piezoelectric transducers ( Figure 11B). This solves the need for chemical fuel of the nanomotor, making the performance of the nanomotor greatly enhanced.
Shafiee and co-workers combined Pt nanomotors with beads that can be detected by the optical system of a mobile phone to achieve a system that can realize rapid detection of Zika virus (≈2 min), with the lowest detection concentration of 1 particle/uL ( Figure 11C). [44] They first cross-linked anti-Zika virus monoclonal antibody (anti-ZIKV mAb) to the surface of PtNPs using a bifunctional cross-linker of 3-(2-pyridyldithio) propionyl hydrazide (PDPH) to fabricate Pt nanomotors, followed by directly coupling the anti-ZIKV mAb to the polystyrene (PS) bead surface using adipic dihydrazide. When Zika virus (ZIKV) is present, they will bind to anti-ZIKV mAb on beads and Pt nanomotors. Pt catalyzes the decomposition of H 2 O 2 to generate bubbles to push the microbeads forward. The mobile phone detects the concentration and movement speed of the beads through optical accessories, and then analyzes the data through the built-in software.
This system has the potential to be widely used in infectious disease diagnosis and treatment monitoring.

pH
pH is an important biomarker that plays a crucial role in numerous physiological and pathological processes. Abnormal pH levels are associated with various diseases such as cancer, diabetes, and cardiovascular diseases, making pH a valuable biomarker for disease diagnosis and monitoring. [76] Recent advances in micro and nanomotor technology have led to the development of more sensitive pH-responsive nanobiosensors that can detect changes in pH levels and provide real-time monitoring, making them promising tools for disease diagnosis and treatment.
Kopelman and co-workers developed magnetic tweezers to assemble swarms of fluorescent chemical sensors containing magnetic components. Magnetic pH-sensitive particles were produced by precipitating iron oxide inside hollow organically modified silica particles and then swelling the shell with a hydrophobic pH-sensitive dye (Figure 12A). [45] These nanoparticles can be controlled by an external magnetic field, and the assembly and sensing of the nanoparticle population can be controlled by a single magnetic tweezer. The luminescence of pH-sensitive dyes is affected by the pH in the environment, and the fluorescence Figure 12. A) Schematic diagram of the structure of the pH sensor. Reproduced with permission [45] Copyright 2007, Elsevier. B) Schematic of microspheres catalyzing the decomposition of H 2 O 2 to move across a pH gradient. Reproduced with permission [46] Copyright 2013, Wiley-VCH. C) Schematic illustration of the motion of PtNP-containing gels in hydrogen peroxide solution. Reproduced with permission [47] Copyright 2016, American Chemical Society. D) (i) Schematic diagram of DNA micromotor fabrication. (ii) DNA nanoswitches hybridize to complementary DNA scaffolds on micromotors. (iii) The pH-dependent triplex-to-duplex transition results in changes in FRET efficiency. Reproduced with permission [48] Copyright 2019, American Chemical Society. emission intensity inside the nanoparticles increases with the pH value. Compared with a single particle, the particle population controlled by magnetic tweezers has higher fluorescence intensity and can be applied to pH detection in various environments.
Chattopadhyay and co-workers found that polymer microspheres containing Pd nanoparticles moved with increasing velocity over a pH gradient after differential catalytic decomposition of aqueous hydrogen peroxide. [46] Nanoparticles can achieve controlled motion when changing their size or applying a pH gradient ( Figure 12B). The detection of pH can be quickly realized by observation under an optical microscope.
In another study, Dong and co-workers fabricated a cassettetype micromotor for pH detection by encapsulating platinum nanoparticles in a gelatin shell. [47] The micromotor is propelled by breaking down the H 2 O 2 in the solution to create bubbles, and the speed of movement increases monotonically with the pH of the analyte solution ( Figure 12C). The main principle is the pH-dependent catalytic activity of Pt NPs caused by the swelling/deswelling of gelatin materials at different pH levels. The detection across the entire pH range can be achieved by observing the motion of this micromotor.
Bio-micromotors propelled by enzymatic reactions produced by imitating biological systems are a relatively new research direction of nanomotors. Sánchez and co-workers functional-ized enzyme-driven hollow silica microcapsules (urease-driven), which moved in the presence of urea. [48] The micromotors were further modified with FRET-labelled pH-responsive DNA nano switches to instantly monitor changes in pH ( Figure 12D). The micromotor can quantitatively detect pH within a few microseconds, enabling dynamic monitoring of pH in the environment.
Our group reported a reversible, pH-responsive motion regulatory mechanism in nanomotors. Succinylated -lactoglobulin and catalase are co-assembled in porous zeolite imidazolate framework-L (ZIF-L) nanoparticles. [49] At neutral pH, thelactoglobulin is permeable, allowing H 2 O 2 to access catalase, which leads to autonomous movement of the micromotors ( Figure 13A). However, at mildly acidic pH, the -lactoglobulin undergoes a reversible gelation process, preventing the access of fuel into the micromotors where the catalase resides. This reversible pH-responsive motility is the first of its kind in chemically driven motors. In one of our following works, we developed a submarine-like micromotor that is capable of directional vertical motion on a centimeter scale by pH-regulated buoyancy control. [50] A pH-responsive, hydrophilic/hydrophobic phaseshifting polymer, poly(2-diisopropylamino) ethyl methacrylate (PDPA), was co-encapsulated with catalase inside ZIF-L frameworks ( Figure 13B). With the aid of PDPA, gas bubbles produced by catalase inside the biocatalytic ZIF-L can be reversibly Figure 13. A) Schematic illustration of the synthesis of pH-responsive biocatalytic micromotors and their pH-controlled motion on/off switch. Reproduced with permission [49] Copyright 2019, Wiley-VCH. B) Schematic illustration of the synthesis of the self-propelled submarine-like micromotors and their pH-controlled vertical motion mechanism. Reproduced with permission [50] Copyright 2019, Elsevier. retained/expelled from the micromotors depending on the pH (the pH switch is at ≈6.4), leading to the buoyancy-controlled ascending or descending vertical motion. Importantly, anti-cancer drug-loaded micromotors showed directional cytotoxicity to 3D cell cultures, depending on the pH of the cellular environment. This study is expected to open new avenues for designing directional propulsion mechanisms for chemical motors, showing potential as autonomous robotics for in vivo delivery in complex biological environments.

Glucose
Glucose is a vital molecule in living organisms and plays a critical role in metabolism, providing energy for various cellular processes. Abnormal glucose levels can indicate a range of health conditions, including diabetes, obesity, and metabolic disorders. Therefore, accurate and reliable glucose detection is essential for disease diagnosis, treatment, and management.
Yin and co-workers fabricated Janus -Fe 2 O 3 /SiO 2 nanoparticles (JFSNs) as miniature biosensors for glucose detection in the blood. [51] Fe 2 O 3 exhibits an intrinsic peroxidase-like catalytic activity and can be used over a wider range of pH and temperatures compared to natural enzymes ( Figure 14A). By immobilizing glucose oxidase (GOx) onto JFSNs, the authors developed a reusable sensor that can detect glucose in complex samples such as serum with high selectivity and acceptable reproducibility. After the de-tection is completed, the particles can be separated and recycled by their magnetic properties.
In another study, Pumera and co-workers used magnesium/platinum (Mg/Pt) Janus micromotors with selfrejuvenating surfaces to improve the electrochemical sensing of glucose in human serum. [52] The glucose detection is based on the glucose oxidase enzyme and ferrocenemethanol shuttle system, and the motion of Mg/Pt Janus micromotors enhances the mass transfer, resulting in increased current response ( Figure 14B). The current signals showed a linear relationship with glucose concentration, and due to the enhanced mass transfer in solution, the detected signal current of glucose in human serum is also enhanced, resulting in lower LOD values.
Very recently, our group fabricated metal-organic framework nanomotors by encapsulating glucose oxidase (GOx) and catalase (CAT) in zeolitic imidazolate framework-8 (ZIF-8) that use biocatalytic cascade induced buoyancy as a driving force and glucose as the fuel. [53] In the presence of glucose, the nanomotors move vertically and collide with the working electrode at the solid/liquid interface, and a current spike due to the collision can be observed ( Figure 14C). It was shown that the number of current spikes (indicates the events of nanomotor as it collides with the electrode) increases linearly with glucose concentration. Beyond glucose sensing, this work offers a chance of exploring the real-time interaction between non-electrocatalytic and redox-inactive particles and electrical signal generation, providing a new dimension to carry out collision experiments. Figure 14. A) Schematic representation of the colorimetric detection of glucose using JFSNs. Reproduced with permission [51] Copyright 2015, American Chemical Society. B) Schematic illustration of the Mg/Pt Janus micromotor-assisted glucose bioassay. Reproduced with permission [52] Copyright 2019, American Chemical Society. C) Collision mechanism of self-propelled nanomotors in the presence of glucose. Reproduced with permission [53] Copyright 2022, Wiley-VCH.

H 2 O 2
H 2 O 2 is a reactive oxygen species that is produced in cells during various biological processes. Its level in biological systems can serve as a biomarker for oxidative stress, which is associated with a range of diseases. [77] Since metals such as Pt can catalyze the decomposition of H 2 O 2 , they are often used to construct nanoswimmers. Crespi and co-workers used Au and Pt to fabricate nanorods that can be driven by the catalytic decomposition of H 2 O 2 at the Pt end. [1] It was found that the movement of nanorods in H 2 O 2 solution was positively correlated with the concentration of H 2 O 2 . However, the high cost of Pt limits their commercial adaptation. Pumera and co-workers used Ag instead of Pt to fabricate nanomotors. [54] In the experiment, it was determined that the concentration of H 2 O 2 controls the moving speed of the Ag catalytic motor, and the shape of the nanomotor also affects its average speed. The nanomotor can move at extremely low H 2 O 2 concentration (0.1%), providing potential application potential for detecting H 2 O 2 concentration in biological samples.

Lipopolysaccharides
Studies have shown that the elevation of lipopolysaccharide (LPS) is associated with poor glycaemic control and subclinical inflammation in diabetes, so LPS has the potential to be a biomarker before inflammation in type 2 diabetes. [78] Escarpa and co-workers fabricated a biosensor for LPS detection using Pt-F 3 O 4 Janus micromotors. [55] Phenylboronic acid (PABA)-modified graphene quantum dots (GQDs) were added during the synthesis of Janus particles (Figure 15). The nanomotor can be driven chemically or magnetically, and the fluorescence of the GQDs is quenched in the presence of LPS in solution that can be observed under a mi- Figure 15. The movement state and fluorescence quenching of the micromotor observed under the microscope before and after adding lipopolysaccharide. Reproduced with permission [55] Copyright 2017, Wiley-VCH.
croscope. This nanomotor can screen complex urine and human serum samples with high selectivity.

Cortisol
Cortisol is a class of steroid hormones, and salivary cortisol is often used as a biomarker of psychological stress. [79] Wang and co-workers fabricated nanomotors that can detect cortisol by naked eye observation. [56] Cortisol-HRP and cortisol compete with antibody sites on micromotors in the TMP/H 2 O 2 system. A blue solution is formed in the presence of Cortisol-HRP. The movement of nanomotors can greatly improve the performance of competitive cortisol immunoassays compared to direct competition assays. This method can detect cortisol at concentrations as low as 0.1 μg L −1 in 2 min. Figure 16. Motion mechanism of electric nanomotors using iodine as fuel. Reproduced with permission [16] Copyright 2011, American Chemical Society.

Iodine
Urinary iodine can be used as a biomarker that directly reflects daily iodine intake to determine children's daily intake consistency with optimal iodine nutrition. [80] The Cu-Pt nanorods fabricated by Sen and co-workers can self-electrophoretically advance in solutions containing iodine. [16] The moving speed of the nanorods varies linearly with the concentration of I 2 in a short time, but further oxidation of Cu is hindered by the CuI layer on the surface of the nanorods in a long time (Figure 16). These nanorods can be used as sensors that detect the concentration of I 2 .

Copper
Copper is an essential trace element in the human body. Changes in the concentration of copper in the human body will have a great impact on the human body. Copper deficiency can cause skeletal defects in young children and osteoporosis in adults; [81] excessive blood copper can induce Wilson's disease, Alzheimer's disease and Parkinson's disease. [82] Wan and co-workers synthesized magnetic mesoporous silica/ZnS·Mn/gold/tetraethylenepentamine/heparin (MMS/ZM/Au/T/Hep) micromotors that can be used to detect and remove Cu 2+ from the blood. [57] The micromotor shows obvious fluorescence quenching after contacting with Cu 2+ , and the intensity decrease is positively correlated with the concentration of Cu 2+ . TEPA, which is linked to the surface of the micromotor by cross-linking with glutaraldehyde, can use abundant amino groups to remove Cu 2+ ( Figure 12A). The results demonstrate that the micromotors can remove 74.1% of blood copper ions due to the synergistic effect of the mesoporous structure, adsorbent functional group, and movement ability. Additionally, the micromotors can selectively monitor blood copper concentration with a limit of detection of 0.33 ppm and a limit of quantification of 1.10 ppm, and a precision (%) of inter-day and intra-day variations of 1.11% and 18.07%, respectively. The presence of Fe 2 O 3 within the micromotor allows the micromotors to be collected magnetically after detection (Figure 17).

Conclusion
In this article, we discuss the common modes of operations of micro and nanomotors and summarize the recent research progress of MNMs in biosensors research. The development of MNMs has gained a lot of attention as promising biosensors due to their unique features, such as their self-propelling ability, fast response time, and ability to detect a wide range of biomolecules. Compared with traditional sensors, the use of nanomotors in biosensors has many advantages. Nanomotors possess the remarkable ability to navigate through a sample, enabling the automatic capture and transportation of target analytes, even within complex environments. This unique characteristic holds immense promise for lab-on-chip diagnosis. The utilization of nanomotors in biosensors brings about autonomous sample manipulation, targeted delivery, enhanced sensor performance, and highthroughput analysis. Consequently, it introduces exciting new opportunities for miniaturization, automation, and enhanced functionality in lab-on-a-chip systems. Nanomotors assume a crucial role in inducing mixing, even within ultrasmall samples, through their active propulsion and stirring of fluids within microscale environments. This active propulsion and the ability to control mixing parameters enable efficient and uniform mixing, effectively overcoming the limitations associated with passive diffusion-based methods. MNMs have the potential to revolutionize the field of diagnostics by providing a faster, more accurate, and more cost-effective alternative to traditional diagnostic methods.
The driving forces behind MNMs can vary widely, ranging from chemical and biochemical reactions to light and magnetic fields. The most used driving force for MNM is hydrogen peroxide, which reacts with the MNM's catalyst to produce a propulsion force. This allows these nanoswimmers to move toward the target biomolecule and bind to it, enabling detection of the target. One of the most significant advantages of using MNMs as biosensors is their ability to actively "find and fetch" targets, thus they can detect low-concentration targets that are difficult to detect using traditional methods. Moreover, the use of nanomotors can reduce the overall cost of diagnostic tests, as they require less equipment and resources than traditional methods. This makes them particularly appealing for use in resource-limited settings, such as developing countries and rural areas.
However, one of the primary limitations of MNMs is their reliance on external driving forces or fuels, such as H 2 O 2 . This presents a significant challenge for in vivo applications, as the requirement for external fuels limits their use in the human body. Hence, one of the primary research foci for nanomotors lies in exploring alternative, more biocompatible fuels that are present in the human body. Several potential fuel sources apart from hydrogen peroxide have been identified and studied, such as urea, [48] oxygen, [52] glucose, [53] amino acids, [83] and reactive metals. [84] However, the utilization of these fuels in nanomotors within the human body is still limited. Future investigations could center on developing nanomotors specifically designed to target and utilize fuels found in specific body regions. Additionally, the use of fuel-free nanomotors presents an alternative and promising solution worth exploring, although actuation of fuel-free MNMs usually requires complex and expensive apparatuses that are difficult to construct. Moreover, external fields such as acoustic and light exhibit limited penetration depth, limiting their applications in biomedicine. Thus, developing more effective fuels or methods to power MNMs is a critical area of research. Another challenge associated with Figure 17. Schematic diagram of the nanomotor for separation and detection of Cu 2+ in blood. Reproduced with permission [57] Copyright 2020, Elsevier.
MNMs is their potential toxicity. The materials used to fabricate MNMs can be toxic to cells and tissues, which can limit their use in clinical settings. Therefore, developing safe and biocompatible nanomotors is a crucial area of research for the development of nanomotor-based biosensors.
Compared to natural motor proteins with high energy efficiency, artificial MNMs have significantly low energy efficiency. The complex biological environment can interfere with propulsion. For example, high ionic strength can cause inefficiency of self-electrophoresis and ionic self-diffusiophoresis, while high viscosity can impede movement. These factors are particularly relevant to biomedical applications since bodily fluids have high ionic strength, viscosity, and flow rates. Future efforts should focus on developing powerful propulsion strategies with high efficiency and stability to overcome environmental interference.
Moreover, for nanomotors to be effective in biosensing applications, it is essential to understand their structure-function relationships. This includes the structure and composition of the nanomotor, the driving mode used to generate motion, and the motion patterns produced by the nanomotor. The composition of the nanomotor can have a significant impact on its function in biosensing. Different materials may be better suited for certain applications, depending on factors such as their biocompatibility, stability, and ability to interact with specific biomarker molecules. By understanding how the composition of the nanomotor affects its function, we can design more effective biosensors for specific applications. The driving mode used to generate motion is also important in biosensing applications. Different driving modes, such as magnetic, acoustic, or chemical, may be more suitable for different applications depending on factors such as the size and shape of the nanomotor, the properties of the analyte and its local environment, and the desired motion patterns. Understanding the driving mode and how it affects the motion of the nanomotor is crucial for designing sensors with the desired properties. Finally, the motion patterns produced by the nanomotor are essential for biosensing applications. Different motion patterns may be more effective for detecting certain biomolecules or for navigating complex biological samples. By understanding the relationship between the structure of the nanomotor and the resulting motion patterns, we can design biosensors with the desired motion characteristics for specific applications.
Despite these challenges, the future of MNMs as biosensors is promising. Ongoing research is focused on developing new types of nanomotors with improved properties, such as higher selectivity, sensitivity, and biocompatibility. Furthermore, researchers are working to develop novel detection strategies that combine nanomotors with other biosensing technologies, such as fluorescence or electrochemical detection, to further enhance their performance. In conclusion, the development of nanoswimmers as biosensors represents a significant advancement in the field of diagnostics. While there are still significant challenges to overcome, the potential benefits of using micro and nanomotors as biosensors, including faster, more accurate, and more costeffective diagnostic tests, make them an exciting area of research with promising prospects.