1 Introduction

The development of rapid and robust diagnostic tools in healthcare systems is essential to reduce disease fatality and enhance patients’ quality of life [1]. In recent years, healthcare has made significant progress in diagnostic technologies and mitigating the impact of diseases on global populations [2]; however, the struggle to sustain the care delivery to a certain group of individuals such as the elderly people persists. The healthcare ecosystem is moving away from large, complex, and stationary diagnostic tools towards small set-ups and devices that can perform multiparameter analysis at the point-of-care to cope with the increasing medical demands of an aging population. From a life science perspective, the miniaturization of diagnostic devices and tools enables innovation in on-site diagnostics, remote patient monitoring, and digital healthcare [3]. Miniature diagnostic tools provide simple, fast, and affordable on-the-spot diagnostic capabilities, making them useful for bedside procedures and field operations [4]. In terms of healthcare practitioners, miniature technologies provide portability and accessibility, overcoming space and resource limitations in healthcare [5].

There are a number of key technologies that enable a seamless transition in healthcare from large set-ups to on-site and mobile health screening services. Magnetic levitation (MagLev) is one such enabling technology gaining increasing adoption for getting close to the end-user and providing more distributed access and personalized care. MagLev introduces magnet-induced density-based analysis for sorting and separation applications [6,7,8]. It is highly suitable for portable, affordable, and fast analysis of biological samples and micro-manipulating (or micro-positioning) of 3D objects. It has great potential to support ongoing miniaturization efforts in clinical laboratory settings [9] and point of care (POC) diagnostics with clinically actionable results, fast turnaround times, ease of use, and cost-effectiveness [10].

MagLev is a relatively new technology that uses magnetic fields to manipulate and position microparticles and cells [11]. When placed between two magnets with opposing poles, an object will rise against the gravitational and buoyancy forces acting on it and levitate to the point where the magnetic and buoyancy forces are in equilibrium [12]. The negative magnetic susceptibility difference between the cells/microparticles and the paramagnetic solution causes cells to levitate at distinct heights within the magnetic field. While the cells tend to levitate towards the point where the magnetic field is the weakest, they remain suspended at a height where the magnetic forces are equal to the buoyancy force of the liquid. This equilibrium height depends on the density and magnetic properties of the cells but not on their size. MagLev is based on the measurement of an intrinsic physical property, density, making it highly applicable to a broad range of density-based analyses [13], measurements [14], separations [15] and classifications [16]. Unlike traditional density-based methods, MagLev offers several advantages, such as high accuracy, low cost, minimal sample volume, and ease of use. Leveraging the direct relationship between an object's density and its response to magnetic fields, it provides a unique and non-invasive approach to the characterization of materials.

The MagLev principle is used in various areas such as the separation of cells and molecules, quality control of materials, and analysis of therapeutic agents [17, 18]. Recent reviews provide a summary of the progress in versatile applications of MagLev systems for readers interested in these aspects [10, 11, 19]. This review summarizes the recent applications of MagLev in the field of diagnostics by analyzing the molecular- and cellular-level changes that accompany the condition of interest (Fig. 1). Our focus is on diagnostic applications of negative magnetophoresis-based MagLev systems, with special attention given to how the advances in other domains such as high throughput screening, automation, and machine learning can help overcome the limitation of existing MagLev systems designed for the analysis of biological samples. Here, negative magnetophoresis refers to the movement of the diamagnetic object toward the minimum magnetic field region in a microcapillary- or microchannel-based platform containing a paramagnetic solution in which the magnetic susceptibility difference (between the object and the environment) is negative [20, 21].

Fig. 1
figure 1

Density-based characterization and separation of cells and molecules using MagLev platforms with different configurations

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2 MagLev platform

Standard MagLev setup consists of a levitation container positioned between two permanent magnets with the same poles facing each other at a distance to create a magnetic field gradient. Levitation of the objects is typically performed in a container filled with a magnetic liquid. The levitation height, which is proportional to the relative density and magnetic properties of the sample and the suspending medium, is visualized with an appropriate imaging system [22,23,24]. In line with the trends to miniaturize components of mechanical, optical, and electronic devices that have gained momentum in the last two decades (miniaturization revolution), the early MagLev systems have evolved into more compact and small setups that incorporate more advanced features [25, 26]. Miniaturization of MagLev systems allowed their integration with advanced imaging techniques, which in return accelerated the use of MagLev technology for bioassays and cellular measurements [12, 27]. System miniaturization offers several advantages for levitating small biological objects such as cells, macromolecules, and their complexes. Despite significant advances in the design and use of the MagLev-based devices, one of the remaining limitations is the prolonged equilibrium time, which varies depending on the size of the object (e.g., the levitation time increases with decreasing object size) due to the Brownian motion [22]. Miniaturization of the MagLev system is an effective approach to overcome the limitation of long levitation time by reducing the distance traveled by small particles and increasing the magnetic field gradient [28]. This compactness and portability of miniaturized MagLev systems allow researchers to conduct experiments in various settings, facilitating on-site experimentation. In addition to the portability, miniaturized MagLev systems require only a small amount of biological samples for each analysis, improving patient convenience and broadening the applications for medical diagnostics and point-of-care testing devices [29]. In this section, the key components of the MagLev system are summarized, together with the theoretical foundations of magnetization behavior and the effects of critical design parameters on the performance of MagLev-based diagnostic tools.

2.1 Theoretical principles of magnetic levitation

The MagLev method is commonly used for the separation of diamagnetic particles or cells [30]. In the MagLev method (Fig. 2), diamagnetic objects spiked in the magnetic medium are levitated under a magnetic field [21]. Magnetic force (\({\overrightarrow{F}}_{mag}\)) and buoyancy force (\({\overrightarrow{F}}_{b}\)) resulting from the acceleration of gravity (\(\overrightarrow{g}\)) and the density difference between the object and the medium are induced on the levitated object. \({\overrightarrow{F}}_{mag}\) is calculated as [22]:

$${\overrightarrow{F}}_{mag}= \frac{({\chi }_{o}- {\chi }_{m})}{{\mu }_{0}} V\left(\overrightarrow{B}\cdot \overrightarrow{\nabla }\right)\overrightarrow{B}$$
(1)

, where \({\chi }_{o}\) and \({\chi }_{m}\) are magnetic susceptibility of object and medium, respectively \({\mu }_{0}\) is the permeability of free space, \(V\) is the volume of the object, \(\overrightarrow{B}\) is applied magnetic induction, and \((\overrightarrow{B}\cdot \overrightarrow{\nabla })\overrightarrow{B}\) is expressed in the three axes (x, y, z) [31]:

$$\left(\overrightarrow{B}\cdot \overrightarrow{\nabla }\right)\overrightarrow{B}=\left[\begin{array}{ccc}{B}_{x}\frac{\partial {B}_{x}}{\partial x}& + {B}_{y}\frac{\partial {B}_{x}}{\partial y}& + {B}_{z}\frac{\partial {B}_{x}}{\partial z}\\ {B}_{x}\frac{\partial {B}_{y}}{\partial x}& {+ B}_{y}\frac{\partial {B}_{y}}{\partial y}& + {B}_{z}\frac{\partial {B}_{y}}{\partial z}\\ {B}_{x}\frac{\partial {B}_{z}}{\partial x}& {+ B}_{y}\frac{\partial {B}_{z}}{\partial y}& {+ B}_{z}\frac{\partial {B}_{z}}{\partial z}\end{array}\right]$$
(2)

\({\overrightarrow{F}}_{b}\) is calculated by [22]

$${\overrightarrow{F}}_{b}=\left({\rho }_{c }-{\rho }_{m}\right)V\overrightarrow{g}$$
(3)

, where \({\rho }_{c}\) and \({\rho }_{m}\) are the density of the cell and medium, respectively. In MagLev, diamagnetic objects move towards low magnetic induction areas, and they are positioned at specific levitation height, where \({\overrightarrow{F}}_{mag}\) is balanced by \({\overrightarrow{F}}_{b}\) [12]:

Fig. 2
figure 2

MagLev principle. The cells with diamagnetic properties levitate at a specific height depending on their density in a paramagnetic medium. During the levitation, the cells move from the area with relatively strong magnetic induction intensity to the weaker region and locate at a specific levitation height, where buoyancy forces are in balance with magnetic forces

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$$\frac{({\chi }_{c}- {\chi }_{m})}{{\mu }_{0}} V\left(\overrightarrow{B}\cdot \overrightarrow{\nabla }\right)\overrightarrow{B}=\left({\rho }_{c }-{\rho }_{m}\right)V\overrightarrow{g}$$
(4)

Under the assumption that the magnetic susceptibility of objects is significantly smaller than that of the magnetic medium, the levitation height of an object depends mainly on the density of objects (but not the volume) within the same MagLev setup, sharing identical medium and magnetic induction characteristics [12]. Objects with higher density experience a greater gravitational force pulling them downward, while low-density objects experience a weaker gravitational force compared to denser ones. Consequently, the density of levitated objects can be estimated based on their respective levitation heights, with higher-density objects are being positioned at lower heights compared to their lower-density counterparts.

The size of objects significantly influences the time required to reach the equilibrium levitation height in the MagLev system. The motion of small objects (< 1 µm) that is governed by thermal (Brownian) mechanics imposes limitations on the precise localization of these objects [11]. To levitate an object, the thermal energy (\({E}_{T}\)) should be less than the sum of the kinetic energy resulting from magnetism and the potential energy resulting from buoyancy. Otherwise, the thermal (Brownian) motion may hinder the levitation of the object in the MagLev system. \({E}_{T}\) is calculated as follows:

$${E}_{T} =kT$$
(5)

, where \(k\) is the Boltzmann constant (1.3806488 × 10–23 m2·kg−2·K−1) and \(T\) is the medium temperature in Kelvin [12]. As the dimensions of particles increase, levitation occurs more rapidly, reducing the analysis time [32]. However, innovative strategies are required to analyze smaller objects (e.g., biomolecules) within a reasonable levitation time. One potential solution to increase the kinetic energy of ultrafine particles is to enhance the magnetic susceptibilities of the magnetic medium [33] and magnetic power of magnets.

2.2 Components

The critical components of magnetic levitation systems include magnets, magnetic medium, sample holder, imaging systems [11].

2.2.1 Magnets

MagLev relies on the interaction of magnetic fields between specially designed magnets and objects. Magnets are used to exert a force that levitates objects/samples against the force of gravity without any physical contact [21]. To date, different magnets have been used for various applications of magnetic fields in transportation [34], particle acceleration [35], healthcare and medical imaging [11], material handling and manufacturing [36], magnetic bearings [37], aerospace and space exploration [38] and entertainment [39]. The magnets used in MagLev-based technologies can be classified as permanent magnets, superconducting magnets, electromagnets, and hybrid magnetic systems [40].

Permanent magnets commonly used in biomedical and healthcare applications operate based on the intrinsic magnetization of certain materials that can maintain a stable magnetic field without an external power source [41]. Permanent magnets are made from hard ferromagnetic materials and several alloys of rare-earth metals that are magnetized during their manufacturing process [42]. They have an inherent magnetic field due to the alignment of the magnetic moments within the material. Hence, permanent magnets are durable and can provide a consistent magnetic field for an extended period [43]. Their compact nature allows them to be used in portable medical MagLev techniques and devices [44]. The most well-known permanent magnets are neodymium iron boron (NdFeB), samarium cobalt, alnico (Aluminum-Nickel–Cobalt), ceramic and ferrite magnets [45]. Magnetization can be increased or manipulated using different magnet configurations such as Halbach array, radial magnetization, multi-pole magnetization, magnetic laminations, or stacking [46]. These configurations can significantly enhance the overall magnetic field strength and/or optimize the magnetic field direction [47]. For instance, in simulations performed with different placements using the Halbach array, magnetic flux densities can be changed and, in this way, magnetic fields in different formations can be created [48]. Another proposed mechanism utilizes a rotating magnetic field generated by a multi-pole Helmholtz coil system to induce active locomotion and targeted drug release [49]. This magnetic torque control mechanism enables precise manipulation of the robot's movement, ensuring controlled drug delivery without leakage. By switching the direction of the thrust force upon reaching the target site, the robot achieves efficient drug release, demonstrating its potential as a functional medical robot for therapy and diagnosis. Additionally, magnetic lamination could be used for magnetic manipulation. The study figured out how the material bends and how waves move through it when soft periodic laminates react to magnetic fields [50]. Such understanding can help predict how stable the material is and how it blocks certain waves in different directions, all by tweaking the magnetic field.

Superconducting magnets play a critical role in the field of MagLev, offering several advantages such as higher magnetic field strength, low energy consumption, reduced heat generation, and improved performance at low temperatures [51]. These magnets operate based on the phenomenon of superconductivity, which allows them to generate extremely strong and stable magnetic fields without the resistive loss of energy [52]. Superconductivity is a quantum mechanical phenomenon where certain materials exhibit zero electrical resistance when cooled to very low temperatures [53]. Superconducting magnets can generate extremely high magnetic fields, which is beneficial for levitating heavy objects and controlling small particles [54]. Once cooled to their superconducting state, they require very little energy to maintain the magnetic field. The most common superconducting materials used in MagLev applications include niobium-titanium (Nb-Ti), niobium-tin (Nb3Sn), high-temperature superconductors (Bi-2212, Bi-2223), and rare-earth barium copper oxides (REBCO) [55, 56].

Electromagnets create magnetic fields through the flow of electric current in a coil of wire [57]. Unlike permanent magnets which are permanently magnetized, an electric current is needed for an electromagnet to generate a magnetic field. The core component of an electromagnet is a coil of wire, typically made from a material with low electrical resistance like copper. When an electric current flows through this coil, it generates a magnetic field around the coil [58]. The strength of the magnetic field produced by an electromagnet is directly proportional to the amount of current passing through the coil and the number of turns in the coil [59]. Magnetism generated by electromagnets can be easily varied in strength and direction, providing precise control in levitation applications [60].

Hybrid magnetic systems of various configurations can be designed by strategically combining different types of magnets and controlling their interactions to achieve customized levitation and manipulation [61], according to the needs of specific application scenarios. They typically combine different magnet types (e.g., superconducting, permanent, and electromagnets) to (1) achieve precise movement of objects or samples, (2) enhance performance, (3) reduce costs, and (4) improve operational flexibility, functionality, and adaptability [62].

2.2.2 Magnetic medium

Materials can be classified into three categories based on their response to magnetic fields: diamagnetic (magnetic susceptibility, \(\chi <\) 0), paramagnetic (0 \(< \chi <\) ~10−2) and ferromagnetic (\(\chi >\) ~10−2, e.g. \(\chi =\) ~105 for iron). Diamagnetic objects with opposing magnetic dipole moment to the applied magnetic field (e.g., majority of cells and proteins) are repulsed from maxima of magnetic field strength [63]. This weak repulsive force under the external magnetic field can be enhanced by increasing the magnetic susceptibility of the medium to manipulate diamagnetic objects. Applications of this negative magnetophoresis-based levitation involve manipulation of the objects using paramagnetic solutions or ferrofluids. The medium to be used in different applications of MagLev is selected based on magnetic susceptibility, compatibility with the sample, transparency, price, and accessibility.

Paramagnetic fluids are generally composed of a paramagnetic metal that produces a magnetic moment due to their unpaired inner-shell electrons such as Manganese (Mn2+) and Gadolinium (Gd3+), and an organic chelating agent or halide [20, 22]. Although aqueous solutions of halide salts such as MnCl2, GdCl3, and DyCl3 are favorable for the levitation of water-insoluble samples, especially owing to their transparency in the visible region, they are not compatible with biological applications due to cytotoxicity [64]. For biological applications, chelate forms of Mn2+ and Gd3+ (e.g., Gd·DTPA, gadolinium(III) diethylenetriaminepentaacetic acid and Mn·EDTA, manganese(II) ethylenediaminetetraacetic acid) are particularly useful due to their biocompatibility [65]. Paramagnetic solutions typically used in the levitation of biological samples involve Gadolinium-based contrast agents (e.g., gadoteridol, gadobutrol) designed for use in magnetic resonance imaging (MRI) [20, 66]. In these structures, the toxicity of free Gadolinium ions is reduced to a biologically safe level by ligand chelation. However, the low magnetic susceptibility of paramagnetic media, especially for the concentrations typically applied in terms of biocompatibility (< 0.2 M), limits their use to levitate small biomolecules [33].

Ferrofluids consisting of nano-sized ferromagnetic particles and a solvent have higher magnetic susceptibility when compared to paramagnetic fluids [67]. Ferrofluids are colloidal suspensions containing monodomain ferromagnetic nanoparticles (usually magnetite, Fe3O4, or maghemite, Fe2O3) with typical dimensions of approximately 10 nm [68]. They are usually coated with surfactants to avoid agglomeration and increase biocompatibility [69]. Over the last decade, significant efforts have been devoted to forming biocompatible ferrofluids for MRI applications, including amino acid-modified magnetic nanoferrofluid [70] and dextran-coated magnetite nanoparticles [71]. Both commercial [72] and custom-made [73] ferrofluids have been successfully applied for the separation of various living cells by MagLev, and their short-term biocompatibility (cell viability > 80% for 1–2 h) has been reported [19]. However, their optical opacity and long-term biocompatibility are still a concern that should be addressed prior to their use in biomedical applications.

Iron oxide nanoparticles that exhibit superparamagnetic properties due to their sufficiently small size (called superparamagnetic iron oxide nanoparticles, SPIONs) have higher positive magnetic susceptibility than paramagnetic materials. Hence, the use of superparamagnetic solutions instead of traditional paramagnetic liquids in MagLev applications accelerates the separation process, particularly when levitating small biological objects [10]. A MagLev protocol using the superparamagnetic solution, commercially known as ferumoxytol (an FDA-approved SPIONs suspension), for levitation of human plasma proteins has been described [33]. Moreover, SPIONs provide faster separation of nanoscale biomolecules due to their high magnetic susceptibility. Hence, SPIONs may enable the levitation of a wide range of biomolecules in a short period of time and contribute to the development of biomedical applications including point-of-care diagnosis and personalized medicine.

2.2.3 Imaging systems

The sensitive imaging of levitated objects within the MagLev system is essential for diagnostic applications [11, 74]. Visualization of MagLev platforms can be performed using various imaging techniques such as visible light (optical) microscopy, infrared imaging, ultrasound, fluorescence imaging, and holographic microscopy [75]. Imaging methods coupled with MagLev platforms differ widely in terms of the amount of detail they provide, sensitivity, resolution, and contrast, and reveal different characteristics of the levitated samples.

Traditional microscopy using visible light is a common method for capturing images of MagLev platforms and providing a visual record of the levitated objects as well as their positions [10]. Visible light microscopy is a standard and widely accessible tool, making it an easy-to-use and cost-effective method for capturing images of levitated objects [76]. It does not expose objects to harmful radiation or high-energy particles, and hence, is considered a safe imaging method for both researchers and samples [77]. However, it may have limited effectiveness for imaging objects with complex internal structures, as it cannot penetrate deeply into objects to reveal their internal composition. Moreover, some materials may not be suitable for visible light excitation, especially if they are transparent or if they do not interact strongly with visible light [78]. The recent advances in MagLev imaging include the development of miniaturized MagLev systems coupled with light microscopy for the separation and density assessment of a diverse array of biomolecules [26], encompassing various cell lines [79], single cell analysis [80], proteins [30], organoids [81], bacteria [82], colonies [83], tissues [84], and polymers [31].

More recently, smartphone-based systems were used for the investigation of biomolecules in specialized MagLev platforms [85,86,87]. These systems can be used for portable and low-cost diagnosis of several diseases by combining the power of microscopic imaging and analysis with smartphone-based methodologies [88]. Smartphone-based imaging systems are also integrated with automated image-processing schemes to perform various tasks such as measuring levitation height, density and concentration [8, 27]. The smartphone-compatible systems have great potential to be used in conjunction with machine learning and deep learning algorithms for automated analysis of the samples within the MagLev system [29].

Lensless holographic microscopy applied to MagLev platforms involves the use of advanced holography techniques to generate real-time 3D holographic images [89]. Holographic microscopy captures the complete 3D structure of levitated objects, providing spatial insights not offered by traditional imaging methods [90, 91]. It enables real-time and dynamic observation of samples and components, making it suitable for studying the density-based, size-based, and other characteristic behaviors of levitating objects under the influence of magnetic fields [92]. While it provides significant advantages such as 3D visualization and real-time observation, it involves complexities in setup and data processing, and requires optically transparent and low-dense samples [93].

Fluorescent imaging is another popular technique often used together with MagLev for the determination and visualization of the levitated samples [94]. Fluorescent markers or dyes can be introduced to samples, enhancing the contrast and visibility of specific elements or materials within the MagLev platform [95]. Additionally, the use of smartphone-integrated fluorescent solutions with MagLev makes such platforms widely applicable to various biomedical analyses [96]. Fluorescence imaging has several advantages relevant to MagLev applications. For example, it can detect and quantify low concentrations of fluorescent substances, providing highly sensitive measurements [97]. It is valuable for biomedical and biological research within the MagLev platform, enabling the study of cellular structures and molecular interactions. However, it may require complex sample preparation steps and high-cost imaging setups [98].

Infrared imaging of MagLev platforms involves the use of specialized cameras or sensors to capture thermal images of the objects or samples within the magnetic field and to identify their characteristics [99, 100]. It is based on the detection of infrared radiation, which is emitted as heat by objects due to their temperature [101]. Infrared imaging in MagLev platforms is a valuable tool for temperature monitoring, early fault detection, and material analysis. However, it has limitations in terms of depth penetration and spatial resolution, and it requires expertise for accurate data interpretation.

Ultrasonic imaging is a non-destructive testing technique that utilizes high-frequency sound waves to provide detailed and real-time information about the condition of materials and structures used in MagLev technology [31]. Ultrasonic technology can be utilized to achieve precise measurements of the levitation heights in the MagLev platform [102]. This technique is non-invasive, portable and versatile, providing real-time imaging and high spatial resolution. However, it has certain disadvantages such as limited penetration depth, low contrast imaging, signal artifacts, and material-specific challenges.

2.2.4 Sample holder

One of the major limitations of a standard MagLev system is its low-throughput nature. The basic MagLev design contains a single sample chamber that can vary in size and shape, such as square cuvettes and microcapillary tubes [12, 66]. Recent modifications in the design of MagLev systems have enabled high-throughput testing of samples, overcoming the limitations of traditional MagLev configurations. For example, a re-engineered configuration, which provides a magnetic field gradient with a NdFeB magnet array compatible with common 96 well plates, has been developed to enable parallel multiple density-based analysis for both biological (human erythrocytes) and non-biological samples [17]. In this design, the levitation in the tubes, each positioned between a pair of same-poles facing magnets, has been imaged with an integrated system, flatbed scanner and simple optical components. The proposed system enables simple and cost-effective high-throughput analysis. A MagLev configuration has also been developed to allow density-based cell analysis under a continuous flow condition within a microfluidic channel, thereby ensuring high-throughput analysis [7].

Several efforts have been devoted to improving the sensitivity and dynamic range of the MagLev systems [21]. The most common approaches involve choosing weaker magnets and reducing the magnetic susceptibility of the medium to increase the sensitivity of the conventional MagLev system. As an alternative strategy, a MagLev design formed by rotating the common configuration to 90° with the magnetization direction perpendicular to the gravitational force has been proposed to increase the sensitivity without destabilization [103]. This rotated configuration of the MagLev system has facilitated high-sensitivity density measurements by using the relatively weak magnetic field gradient parallel to the faces of the magnets positioned opposite each other. The sensitivity of the MagLev system in a rotated configuration can be enhanced by reducing the magnetic gradient along a selected axis, through elongating the magnets along that axis or by increasing the distance between the magnets. A high distance between magnets (\(d\) = 18.9 cm) has enabled an enhancement of the resolution up to 100-fold over the classical configuration (down to \({\Delta \rho }_{min}\) = 2 × 10−6 g cm−3). Unlike the standard configuration, the rotated configuration offers another advantage as the open side of the levitation chamber remains unobstructed by a magnet, allowing for straightforward addition or removal of sample and medium, and adjustment of medium properties without disturbing the chamber. This feature enables the translation of this technique into high-throughput industrial settings. Another MagLev configuration has been developed to increase the dynamic range of the density by rotating the standard configuration, which measures in a narrow density range (0.8 g cm−3 < \(\rho\)  < 2.3 g cm−3), with the angle θ (0°< \(\theta\)  < 90°) relative to the gravitational axis [104]. This modification has decreased the gravitational acceleration along the direction of measurement, and therefore, increased the range of densities that can be levitated [104]. The system greatly broadens the spectrum of measurable densities in standard MagLev setup, allowing for measurements ranging from ~ 0 g cm−3 to ~ 23 g cm−3. A rotation-based MagLev design that uses centrifugal acceleration has also been presented to dynamically tune the sensitivity and density range [105]. In the static MagLev design, objects with similar density may cluster along the centerline of the device and the resolution may be insufficient for measurement. In this platform, the MagLev device is fixed eccentrically on a rotating disk, and the gradual increase in the rotating speed deviates the levitated samples from their original position. The sensitivity and the resolution of the measurement can be tuned by adjusting the rotating speed. In the proposed design, the levitation chamber is positioned between four identical magnets, two of which are arranged coaxially with same-poles facing to focus cells along axes where both gravitational and centrifugal forces are effective. The sensitivity of the measurement can be improved by adjusting the eccentric distance or rotational speed.

3 Analysis of biological materials

MagLev technology facilitates the development of various methods for biological analysis due to its label-free, contact-free and high-sensitivity nature, holding a great potential for applications across healthcare and the pharmaceutical industry. The versatile nature of MagLev stems from the intrinsic differences in density and magnetic properties among biological entities. This section describes applications of MagLev for cellular and molecular analysis, and separation of various samples, including cancer cells, blood cells, bacteria, protein corona structures, antigen–antibody complexes, and even drugs (Fig. 3). A summary of MagLev-based analysis methods for cellular and molecular applications is provided in Tables 1 and 2.

Fig. 3
figure 3

Illustrations of MagLev-based platforms used for diagnostics. A) Cellular level analysis. (i) Detection of polymorphonuclear leukocytes (PMNs) activation based on PMN density change (activated PMN in red and resting PMN in blue) [66]. (ii) Separation of particles and circulating tumor cells (CTCs) (from blood cells) according to their density in a smartphone-based cell separation setup [85]. (iii) Sorting of CTCs and white blood cells (WBCs) on the MagLev platform. Cells are dragged by a flow (Q) while being lifted to a constant height depending on their density under buoyancy and magnetic forces. Lower-density cells (CTCs) are collected from the upper outlet, while higher-density cells (WBCs) are directed to the lower outlet. g is the gravitational acceleration and B is the magnetic induction [7]. (iv) Levitation of uninfected red blood cells (uRBCs), ring-stage synchronized infected red blood cells (ring iRBCs), and infected red blood cells (mature-stage iRBCs), in low-density medium (left) and high-density medium (right) [106]. (v) General flow of the airborne virus detection including collection of air, concentration of cells on the MagLev platform, extraction of virus and then extraction of RNA, PCR process, and identification of viruses [107]. (vi) Use of magnet arrays to enable high-throughput density-dependent measurement in MagLev [17]. (vii) Determination of density and size of different cell types in a pump-free automated microfluidic MagLev system between two ring magnets [108]. B) Molecular level analysis. (i) Illustration of the sandwich immunoassay with antigen Interleukin 6 (IL-6), high-density antibody-coated beads (1.2 g cm−3), and low-density antibody-coated beads (1.05 g cm−3). Separation of low-density antibody-coated beads, high-density antibody-coated beads, and aggregated beads IL-6 on the MagLev platform [109]. (ii) Detection of the change in density of polystyrene nanoparticles (NPs) and protein corona (PC)-coated polystyrene NPs on the MagLev platform [110]. (iii) Protein detection via the MagLev platform. The method involves the use of polymeric microspheres as mobile carrier surfaces and magnetic NPs as labels. Target proteins are captured on the polymer microspheres and magnetic NPs are then attached to this microsphere-protein complex, resulting in a significant difference in the magnetic properties of the polymer microspheres compared to protein-free polymer microspheres. With the change in magnetic properties, a difference occurs in the levitation heights based on the amount of target proteins [30]. (iv) Separation of enantiopure crystals (S-ibuprofen, in red) and ibuprofen racemic crystals (RS-ibuprofen, in green) based on their densities using MagLev platform [111]. (v) MagLev NP-assisted blood (NEB) test for cancer detection. Human plasma is separated from blood collected from non-oncological patients and patients affected by different types of cancer. Then, plasma incubated into NPs solution such as graphene oxide (GO) solution and labeled with GO. Profiles of GO-plasma complexes are measured on the MagLev platform [112]. (vi) Separation of dilute fentanyl in the mixture of heroin and diluent (α-lactose) in a MagLev platform. Demonstration of the decisive difference in the peaks obtained by Fourier transform infrared spectroscopy – attenuated total reflectance (FTIR-ATR) as a result of analyzing the samples taken from the lower and upper levels in the MagLev platform [113]

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Table 1 A summary of MagLev-based analysis methods for cellular applications
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Table 2 A summary of MagLev-based analysis methods for molecular applications
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3.1 Cellular level

MagLev platforms offer a unique approach for cell-based assays, providing label-free, high-resolution, and density-based separation and analysis. For instance, the MagLev platform integrated with a mobile phone was used for the detection of sickle cell phenotypic [27]. In order to provide a deoxygenated environment, blood samples from sickle cell (SS genotype) patients have been prepared in a paramagnetic medium using sodium metabisulfite. The samples were subsequently introduced into the capillary and placed between N52-grade neodymium magnets. After waiting 10 min for the equilibrium, images were taken with a mobile phone camera using an application interface developed specifically for the application. Sickle RBCs have higher density than healthy RBCs, and therefore, have lower levitation height, which was captured by the platform quickly using a very small sample volume. A semi-automated and handheld iteration of the MagLev platform was also used with a step-by-step guide for the density analysis of sickle cells [114]. On the MagLev platform, RBCs containing sickle and normal hemoglobin (HbS and HbA) are analyzed according to their density profiles. Since RBCs with sickle hemoglobin are denser, low levitation height was observed. The individual RBC levitation density profile enables RBC detection within minutes with a resolution of up to 0.0001 g mL−1 [25].

The versatility of the MagLev technology enables their adaptation to high-throughput systems. For density-based measurement in 96 wells, a MagLev platform was able to perform high-throughput analysis of both biological and non-biological materials [17]. The workflow involves preparing various samples with paramagnetic solutions (MnCl2 for non-biological samples and paramagnetic Gadavist for biological samples) and loading them into individual 96-well plate tubes placed between parallel long magnets. On the platform, the densities of RBCs, copper powder, polymer particles, cholesterol crystals and chlorotoluene as a hydrophobic liquid were measured. The multi-magnet array's augmented magnetic strength enables the use of low concentrations of paramagnetic solutions, offering a significant advantage for biocompatible applications.

Expanding beyond the RBCs, MagLev-based cytometry was also shown to be applicable for white blood cell (WBC) analysis [8]. Compared to RBCs, WBCs displayed higher levitation heights in the MagLev platform. Another MagLev platform iteration, called i-LEV, has smartphone integration for the analysis of cells in the blood sample [86]. This platform allows the analysis and categorization of WBC and RBC based on their levitation height profile. Using i-LEV, they demonstrated the significant impact of the cell numbers on both the bandwidth of blood cells and the time required to reach equilibrium. In another MagLev-based smartphone device, successful detection and sorting of blood cells and polystyrene microparticles (0.97 g cm−3) from large sample volumes under a flow were shown [6]. The system can be controlled with an Android application, providing real-time monitoring and sorting of particles.

A MagLev platform has been developed to quantify the morphology and size of WBCs [115]. In the proposed approach, blood cells were loaded into the microcapillary with 21 mM Gadavist solution which reached levitation height after 20 min. Their results indicated that the size parameters of leukocytes in the blood taken from septic patients and healthy samples were significantly different. In another study, a MagLev platform was employed to investigate the cellular effects of intravenous (IV) fluids commonly used in healthcare [116]. The results indicated that exposure to IV fluids could differently affect the viability and morphological properties of endothelial cells and monocytes. Particularly, the observed decrease in levitation height, indicating an increase in cell density, suggested that dextrose treatment led to cell death or damage.

Extracellular matrix (ECM) stiffness is a key feature associated with cancer progression and metastasis. Mechanical characteristics of ECM can potentially affect the physical properties of cells [130, 131] including density [28]. To investigate the relationship between ECM stiffness level and single-cell density, levitation profiles of MDA-MB-231 breast cancer and A549 lung cancer cells cultured on varying collagen densities were compared [28]. When the cells were cultured on different concentrations of collagen (0.36, 0.72, and 1.44 mg mL−1) for 7 days, the density distribution of breast and lung cancer cells exhibited changes in single cell density variance, albeit no observed change in average densities. Furthermore, researchers explored the impact of inhibiting protein translation on cell density using Salubrinal, a phosphatase enzyme inhibitor, and demonstrated a decrease in cell density. Also, when the cells were cultured with increasing collagen density, they revealed that the expression of proteins related to mechanosensing processes was changed. While the direct influence of fiber density on biological characteristics of cancer cells was not specifically investigated in the study, a discernible biological effect and alterations in the range of cell density were evident.

Single cell separation through Maglev offers applications for circulating tumor cell identification. Using the density-based MagLev system, a significant difference in density between cancer cells and blood cells was revealed [12, 85]. For instance, the density of various cancer cell types (including breast, esophageal, lung, and colorectal cancer) was lower compared to healthy RBCs and WBCs [12]. Also, the density profile of the cells indicated that breast cancer cells (MDA-MB-231) constituted the most heterogeneous cell population, exhibiting a wide range of density among individual cells in the population. This density difference between different cell types can be used for diagnostic purposes. For cells with similar densities, auxiliary-integrated magnetic levitation systems can be used to capture small differences in density and to distinguish cell types in a heterogeneous solution. One such example is magnetic levitation-based cytometry, capable of contrasting blood cell populations like sickle RBCs, activated PMNs, CTCs, and even differentiating young and old RBCs [66]. The analysis started with preparing samples in a 30–50 mM paramagnetic medium (Gadavist) and loading them into a microcapillary positioned between magnets. Over 10–20 min, cells reached a levitation equilibrium, revealing distinct levitation heights for different cell types. For instance, young RBCs with lower density (~ 1.09 g mL−1) levitated to higher positions compared to old RBCs (~ 1.11 g mL−1). This principle has also been used for PMNs, which activated cells showing altered intracellular components and different magnetic properties. The platform also enabled the visualization of the interaction of PMNs with bacteria like Salmonella based on unique levitation profiles. Furthermore, they were able to differentiate CTCs from PMNs as well as healthy RBCs from RBCs from sickle cell anemia, highlighting the diagnostic potential of the MagLev platform. In a separate study, a hybrid approach was employed by combining the MagLev system with the deep learning-based object detector YOLO to identify individual cells within heterogeneous cell populations closely positioned due to their similar densities [117]. When the breast cancer cells and monocytes were mixed and levitated, the hybrid MagLev system demonstrated high identification efficiency, surpassing 85% for breast cancer cells and exceeding 90% for monocytes. Also, an automated image analysis system named Fastcount was developed for quantification and characterization of CTCs and circulating tumor cell clusters (CTCCs). This MATLAB-based algorithm was employed to analyze immunostained CTCs and CTCCs separated from blood cells using the MagLev system. It was reported that the Fastcount algorithm enabled counting 120 times faster with low deviation compared to manual counting [118].

While current MagLev cytometry setups mostly involve a capillary channel between two magnets, an alternative pump-free microfluidic MagLev system consisting of a chamber connected to five exhaust outlets between two ring magnets was proposed for sample loading and measurement [108]. This system enabled the automatic fluid movement into the measurement chamber with smaller sample volumes, approximately 4 µL, in 16 s. The cells in the paramagnetic solution were levitated and formed specific bands in the measurement chamber according to their density. Using this technology, the single cell density and size of different cell types including various cancer cell lines (HCT116, HeLa, HT1080, and Huh7) and retinal pigment epithelial cells (ARPE-19) were determined. During levitation, researchers observed morphological changes in cells from round and transparent to rough and fuzzy because fetal bovine serum used as levitation solution included biomolecules attaching to the cell wall without affecting the levitation profile of the cells in the system. They demonstrate that the cancer cells exhibited a more varied range of density and size. Also, the drug response of A549 lung cancer cells to Gefitinib was observed. After treatment with high concentrations of Gefitinib, the density of lung cancer cells was decreased, forming a heterogeneous cell population with different densities.

In another study, the MagLev system was developed for real-time detection and separation of cancer and blood cells by integrating a continuous-flow MagLev system with a smartphone, designed for use as a diagnostic tool in resource-limited settings [85]. The smartphone-based MagLev device was manufactured using three different 3D printing methods: polyjet, stereolithography, and fused deposition modeling. The study evaluated the printing time, cost, and performance of five critical device components for comparison. Polyjet 3D printing, despite being more expensive, produced all working parts with greater accuracy and precision with the most reliable mechanical properties compared to alternative printing methods. The study also demonstrated the relationship between flow rate and separation efficiency. A low flow rate of 0.006 mL min−1 outperformed higher flow rates (0.012 mL min−1 and 0.06 mL min−1) for separation, based on the increased exposure time to the applied magnetic field. Using the fabricated device, breast, lung, ovarian, and prostate cancer cells were separated from blood cells with a separation distance of around 100 μm. Another study further advanced continuous-flow MagLev technology by developing a microfluidic platform with a single inlet, two outlets, and a centrally positioned microseparator [7]. Magnetically-guided cells were directed to either top (lower density) or bottom (higher density) channels through the junction formed by the microseparator. The study identified the parameters that affect sorting efficiency, including the flow rate, cell density, and size, the concentration of the paramagnetic medium, and the position of the microseparator. The system successfully sorted MDA-MB-231 cancer cells from a mixture with U937 blood cells after the optimization of separation parameters. The use of a 5 mM paramagnetic medium and a flow rate of 1 mL h−1 yielded a low sorting efficiency (< 40%) of cancer cells, which was increased to approximately 70% upon raising the paramagnetic medium concentration to 15 mM. Moreover, the separated cells exhibited high viability. The developed technique can also distinguish minor density differences and separate rare subpopulations of cells (e.g., circulating tumor cells constituting 0.01% of the entire population).

The versatility of MagLev technology allowed applications for the diagnosis of infections caused by microorganisms and viruses. In a study, the detection of RBCs infected with malaria parasites was investigated based on the changes in the magnetic susceptibility and single RBC density [106]. The study first examined the individual effects of magnetic susceptibility and density on the levitation profile. It was observed that paramagnetic cells treated with NaNO2 levitated less than diamagnetic cells in a low-density medium, but there was no significant difference in a high-density medium. Examining cells with different densities obtained by altering the medium tonicity revealed that cells with lower density (in a hypotonic environment) were positioned higher. When the researchers levitated malaria-infected cells, leading to changes in both density and magnetic susceptibility parameters simultaneously, they achieved better separation in a high-density medium compared to a low-density environment due to the combined effect of density and magnetic susceptibility, thus improving resolution.

Another study introduced a non-invasive system using MagLev for isolating and purifying airborne viruses from indoor environments [107]. The system utilized a standard MagLev platform to levitate airborne viruses at a specific levitation height based on their density within a cuvette. The study tested the MagLev system's efficacy in levitating various types of microorganisms, including bacteria (E. coli), bacteriophages (MS2), and human viruses (SARS-CoV-2, influenza A), both in pure samples and samples from complex air environments. Following levitation, samples were collected from four distinct fractions within the MagLev column, and qPCR and colorimetric assays were used to detect the presence of target organisms in the samples. The results indicated that the bacteriophages and viruses were primarily enriched in fraction A, the highest level in the levitation column. Moreover, air samples containing different concentrations of heat-inactivated SARS-CoV-2 also yielded similar results, with the highest viral RNA copies consistently detected in the fraction. Notably, RT-qPCR results demonstrated that the detection rate of viruses in air samples was higher with the MagLev system than without, suggesting that the system effectively enriches and purifies the samples. However, the study also acknowledged its inability to discriminate between viruses with similar densities.

The inherent density of the cells can be altered by internal lipid content, a characteristic affected by various diseases like neutral lipid storage disease, non-alcoholic fatty liver disease, and obesity. A decrease in cell density due to increased lipid accumulation through the process of adipogenesis of bone marrow cell lines and lung cancer cells was determined by the MagLev system [89, 119,120,121]. Also, in another study, the MagLev platform was utilized to sort cardiomyocytes suffering from abnormal lipid storage due to adipocyte triglyceride lipase deficiency [26]. The levitation profile of healthy and diseased human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) indicated that the diseased cells were levitated at higher levels due to decreased density resulting from abnormal lipid accumulation. Thereby, diseased cells were collected from the top outlet, whereas healthy cells were sorted from the bottom outlet. The post-sorting analyses revealed that the MagLev-based sorting system did not cause any impairment of the sarcomeric alignment, or contractility function of CMs. On the other hand, researchers demonstrated impaired sarcomeric alignment in diseased CMs compared to healthy ones.

Evaluation of biological samples in forensic cases can benefit from MagLev-detected density differences. In an example study, sperm cells were separated from epithelial cells with MagLev for subsequent genetic analyses [122]. The flow-based magnetic levitation system was optimized using various concentrations of paramagnetic agents and cell mixtures including dilutions of sperm cells spiked in a much denser population of epithelial cells due to rare sperm populations in samples. During co-levitation of the cells, sperm cells were located at higher levels in the channel with an equilibrium time of < 10 min, and better separation was observed at lower paramagnetic medium concentrations (30 mM). The system was further tested on mock forensic samples. Although the forensic samples mostly consisted of dead cells or cells with compromised structural integrity, the MagLev system achieved over 90% sorting efficiency and more than 97% purity in the sorting process.

Dead cells in cell cultures can compromise drug screening, functional studies, and data quality, as well as the success of cell therapy and tissue engineering studies [132,133,134,135]. Therefore, the separation of dead and live cells is fundamental for maintaining the integrity of experiments, ensuring the reliability and repeatability of results, and supporting various applications in research. Cell death is one of the biological processes that can alter the density of the cells [12, 133]; thus, the MagLev system offers the capability to separate live and dead cells based on their density. Through drug and chemical treatments of cells, studies documented that dead cells had higher density compared to live cells, as well as wider density ranges compared to healthy cells [12, 89]. Moreover, in the study, the MagLev system was combined with lensless holographic microscopy to analyze cell densities using an imaging sensor [89]. Thereby, the system called HologLev also offered the opportunity for highly efficient and automated cell analysis with on-site monitoring in an incubator. Furthermore, a novel application of MagLev has been reported a rapid and label-free sorting of live and dead cells in a microfluidic system [123]. The authors successfully enriched viable breast cancer cells from heterogeneous mixtures with high efficiency and purity by capitalizing on the differences in cell density between live and dead cells. Using input purities ranging from 10 to 50%, they achieved high output purities of up to 80% within a 30 min operation.

3.2 Molecular level

MagLev method provides density-based analysis and detection of biomolecules such as proteins, antigens, and antibodies [21]. In a study, a magnetic-linked immunosorbent assay (MeLISSA) was developed for detection of cell membrane-bound and soluble antigens. The cell-bound or soluble target antigens were levitated with antibody-coated beads of different densities, shapes, or colors in the capillary tube containing a 40 mM paramagnetic medium. After the interaction between antibody and antigen, the complex levitated between noninteracting beads and target antibodies based on the new density [109]. Similarly, in another study, density-linked immunosorbent assay (DeLISA) was developed using metal-amplified density assay (MADA) which is a quantitative immunoassay method using MagLev in order to detect antibodies against Hepatitis C virus (HCV) NS3 protein and T. pallidum p47 protein [128]. Using anti-HCV NS3 antibody functionalized microsensor beads (MB), HCV was detected by smartphone-assisted MagLev platform. The HCV NS3 protein was captured by MBs at the 50 mM paramagnetic medium concentration. The protein detection was determined by decreasing the levitation heights of MBs due to the increase in the MBs density [88]. HCV was successfully detected with 50 µg mL−1 limit of detection (LOD) value. With the same magnetic levitation platform, Bovine Serum Albumin (BSA) was also detected with antibody-functionalized polystyrene microspheres [129]. LOD value was calculated as 4.1 ng mL−1.

In MagLev setup, polymeric microspheres were used as mobile assay surfaces, whereas magnetic NPs were used as labels [30]. The technique employed the difference in the levitation height of protein-microsphere-NP complex compared to that of polymeric microspheres (without target proteins). In this assay, the magnetic susceptibility of microspheres was increased in the presence of target proteins, with magnetic NPs specifically attached to them. This resulted in a decrease in the levitation height of the microspheres. The detection limit of this method was ~ 110 fg mL−1 for biotinylated BSA (b-BSA). They also tested the designed MagLev system for monitoring mouse immunoglobulin G (IgG) and achieved detection limits of 1.5 ng mL−1 and > 10 ng mL−1 in buffer and serum, respectively.

Biomolecule mixtures can be levitated with MagLev setups involving SPION suspensions due to their high magnetic susceptibilities [33]. This technique was used for the analysis of disease-based expressions of important proteins in plasma (~ 1.03 g cm−3) [16]. In this study, diagnostic analysis was performed according to the differences in the expressions of the proteins. The samples prepared with SPIONs were loaded into the cuvette and equilibrated between magnets for 180 min. Then, a histogram curve was created based on the differences in levitation heights extracted from images using machine learning algorithms. Levitated plasma proteins were collected and molecular analysis (LC–MS/MS) was performed to show the spectrum difference of samples collected from the MagLev system of healthy individuals and patients using opioids. It was observed that the bands formed at the top of the sample holder contained more proteins in the samples of patients with opioid disorder, and proteins that were known to be at high levels in plasma during opioid use were also abundant in these bands. This suggests that the developed system can be a useful tool for the observation and identification of proteins.

MagLev platforms have been used for the detection of various early-stage tumors based on protein biomarkers. One promising approach for early cancer diagnosis emerges from the identification of the protein cloud (so-called protein corona) that surrounds NPs upon introduction to blood samples from cancer patients. MagLev profiles of protein-coated NPs were shown to contain potential cancer markers [112, 136,137,138]. A NP-enabled blood (NEB) test has been developed by indirectly characterizing personalized PC using MagLev [112]. This test was based on the analysis of the levitation profiles of PC-coated GO NPs. Results showed strong cancer type-dependent specificity and sensitivity, with the best identification rate being obtained for breast cancer and pancreatic ductal adenocarcinoma (PDAC). In the follow-up study, they tested blood-based biomarkers using corona-coated GO NPs for early detection of PDAC and they managed to distinguish PDAC patients from healthy ones with an accuracy of 91% [125]. Another attempt to detect PDAC by levitating GO NPs decorated with human plasma proteins coronas, using blood samples collected from both PDAC and non-oncological patients (NOPs) [124]. They managed to distinguish PDAC patients from NOPs with a specificity and sensitivity of 80% and 100% respectively, reaching 90% classification accuracy. To further increase the accuracy of MagLev-based diagnostic devices employing NP-PC complex as a biomarker, the influence of system parameters were assessed such as total flow rate and flow rate ratio on the MagLev profile of GO and plasma protein complex generated in a microfluidic environment [126]. In another study combining PC-attached nanostructures with a MagLev-based approach for PDAC detection, a success rate of ∼95% was achieved by coupling common inflammatory biomarkers with levitation profiles of PC-coated GO nanosheets [127]. MagLev systems can also provide information about the identity and heterogeneity of PC formed around NPs upon exposure to biological fluids and can allow monitoring the evolution of the NP-PC that is crucial for understanding their in vivo responses, such as bioactivity and safety [110, 139].

Although material characterization, forensic chemical analysis, and separation of crystalline mixtures in different forms are challenging processes, MagLev appears as an alternative to existing systems used in the examination of these structures. For example, a 30° tilted MagLev platform was developed for enantiomeric purity analysis of Ibuprofen [111]. The samples loaded in a glass bottle in 0.55 M MnCl2 paramagnetic solution reached equilibrium height. The sample mixture prepared in crystal forms as S and RS was exposed to a magnetic field for 12 h. The two populations that came to equilibrium were then collected and their purity was determined in differential scanning calorimetry (DSC) analysis. MagLev provided an alternative crystallization methodology, separating enantiomer mixtures with 95.1% purity, proving that it could be an effective, easy, and portable system in the pharmaceutical industry. In a study conducted to increase the sensitivity of MagLev, polymer and drug spheres were analyzed with precision according to their density under a magnetic field [103]. At these rotated MagLev platforms, magnets are placed in a way that the gravitational force is perpendicular to the magnetic force. This configuration has increased separation sensitivity, providing 100 times more sensitive analysis compared to standard platforms. The drug capsule was opened and the density distribution of the microspheres inside was observed in the developed rotated MagLev for drug quality control. Illicit drugs were also analyzed using the MagLev technique [113]. The analysis was carried out in a cuvette placed between magnets. The sample mixture spiked in paramagnetic solution was prepared in 23% hexane and 77% tetrachlorethylene by volume to prevent the dissolution of illicit drugs and then it was loaded into the cuvette for inspection. In this way, it has been shown that mixtures of illicit drugs with different specific densities (0.66–1.77 g mL−1) can be distinguished without requiring additional analysis at low sample concentrations. The separated drugs in the MagLev platform can be collected with a Pasteur pipette for more detailed assessments with molecular methods, such as FTIR-ATR, and NMR.

4 Challenges and emerging technologies

MagLev technology has emerged as an innovative technique for disease diagnosis and biomarker discovery based on magnetic field-induced changes in density. The maturation of flow-based microfluidic systems has revolutionized its capabilities, enabling efficient and cost-effective analysis of larger sample volumes and large-scale applications. While early applications of MagLev in disease diagnostics relied mainly on circulating cells and cancer cells, recent research demonstrated the great potential of MagLev systems in detecting a variety of diseases such as neutral lipid storage disease, Hepatitis C, and other infectious diseases. One of the key challenges in MagLev-based diagnostics is the difficulty in achieving high resolutions that would allow the detection of even subtle density differences. For instance, when used for testing biological air pollution in indoor environments, the MagLev system can be inadequate in distinguishing different types of viruses due to similar densities of strains [107]. In such cases, hybrid systems based on deep learning and artificial intelligence may be applicable to enhance the recognition of distinct structural features in target cells and the precision of density-based detection [117].

Miniaturized diagnostics encompass methodologies and devices that have been downscaled in size, often employing microfluidics, nanotechnology, and sensor technology, to facilitate cost-effective, portable, rapid, and accurate analysis of biological specimens [140]. These compact diagnostic platforms offer numerous advantages over traditional diagnostic modalities, including reduced sample requirements, accelerated testing times, and suitability for decentralized healthcare settings [141]. These miniaturized diagnostic tools serve for a broad spectrum of medical purposes, targeting various pathogens such as bacteria, viruses, and fungi, markers for genetic disorders, or other analytes indicative of specific conditions [142,143,144,145,146,147]. However, the advancement and commercialization of these tools may face several challenges, including complex design schemes [148, 149], principles of labelled detection [150], costly detection methodologies [140, 151], and complex operational principles [152, 153]. In contrast, MagLev platform operates on a versatile principle, enabling the analysis of various targets, such as whole cells or specific proteins, with or without labels, facilitating the diagnosis of diverse diseases [26, 88, 129]. Moreover, it eliminates the necessity for complex detection principles commonly present in other systems by using simple optical imaging techniques, including smartphone-assisted imaging [27, 88]. Its user-friendly nature, cost-effectiveness, and accuracy hold promise for extending the applications of this technology as diagnostic tools in resource-limited settings [25, 30].

Traditional analytical methods, despite offering multiple tests for diverse biological fluids (e.g., blood), still face significant challenges in detecting low-concentration targets such as CTCs and hematopoietic cells within the sample [7, 154,155,156]. The ability to detect low concentrations provides novel insights into biological processes and disease states by allowing the analysis of the cell's heterogeneity as well as its interactions with other cellular components [157, 158]. Additionally, conventional methods often mask rare cells like CTCs because of the overwhelming number of blood cells (WBCs, RBCs) in bulk analysis [159]. Maintaining a consistently high-yield processing for reliable single-cell analysis presents a challenge for conventional methods such as Ficoll-Paque or Onco-Quick, primarily due to their reliance on bench-top devices, impacting practicality, cost, and overall efficiency [160]. However, these methods are limited by the bench-top devices that affect the practicality, cost, and yield. At this point, MagLev technology emerges as a promising solution for single-cell analysis. Its unique ability to measure cell densities label-free at the single-cell level, offers distinct advantages over conventional methods [11]. Separation, analysis, and combination of biological samples according to cell densities performed in MagLev are of critical importance in single cell analysis [7, 84, 89].

Although the MagLev platform works with density differences of objects to produce results, process time that is related to the system’s equilibrium depends on the size of particles or cells. For example, particles having nearly 1 µm size can be affected by Brownian motion in MagLev and hence, they cannot be levitated at a certain height [22]. To overcome these limitations, magnetic field can be increased with stronger magnets [161]. Moreover, either the concentration of the paramagnetic solution can be increased or a magnetic medium with improved magnetic susceptibility can be used. However, the biocompatibility of the chosen magnetic field and paramagnetic solution should be taken into consideration to preserve the integrity or the viability of the sample of interest [12].

To date, molecular analysis studies using MagLev technologies have focused on the identification of early-stage tumors and protein biomarkers The detection capacity of MagLev-enabled molecular-level detection and identification systems can be improved by optimizing the concentration of paramagnetic solution, type of nanoparticles, and exposure time to samples. Modification of micro/nanoparticle assemblies in terms of core and surface composition could further improve diagnostic testing sensitivity. For example, microparticles could be labeled with higher magnetic susceptibility NPs for more sensitive protein detection. The sensitivity could be further improved by modifying micro/nanoparticle surfaces to reduce nonspecific adsorption in plasma [30]. Protein detection assays could be adopted for the detection of other biomolecules by changing the recognition part on the particles. Moreover, NEB tests hold great promise for identifying proteins (or the changes in protein structures) with PC-coated NPs that are predictors of clinical outcomes or responses to treatment. Such innovative approaches can provide a reduction in costs and turnaround time in clinical settings, contributing to the affordability and sustainability of healthcare systems.

MagLev systems hold significant potential for to determine the biophysical properties of sub-cellular vesicles. While there is limited evidence-based information for the direct utilization of sub-cellular organelle densities as diagnostic markers, it is known that intracellular biophysical properties are influenced by biological states, such as disease states and physiological conditions [162, 163]. With advances in complementary technologies that improve the detection range and precision of MagLev systems, it may become feasible to track various pathophysiological changes through levitation of small biological particles like mitochondria in the near future.

The development of hybrid systems has opened an innovative window for classifying and sorting cells [164,165,166,167]. For instance, using magnetic, gravitational, and drag forces for sorting of cells in a MagLev setup, the separation efficiency of cancer cells has been increased by up to 70% [7]. Also, polymer particles with densities of 1.02 g mL−1 and 1.09 g mL−1 are separated from each other with ~ 90% efficiency. Moreover, the integration of morphological analysis with MagLev techniques can boost the performance of cell analysis [116].

MagLev systems can be harnessed to create highly precise and versatile platforms using machine learning-based technologies for the study of biological molecules, cells, and tissues [117, 168, 169]. Machine learning algorithms can analyze the levitation data to detect the presence of biosamples with high sensitivity, offering a non-invasive and early diagnostic tool [170]. Machine learning-based technologies integrated with MagLev systems offer a powerful combination of precise manipulation, non-invasive analysis, high-throughput and label-free measurements, enabling advanced research and applications in life sciences [116]. For this purpose, after obtaining morphological image data from biosamples, tracking, variability analysis, segmentation, classification, and automated analysis can be performed with machine learning techniques [171]. Besides imaging, machine learning offers several advantages as predictive modeling, optimization, automation and control, real-time monitoring, and analyzing [171,172,173,174]. Machine learning algorithms such as support vector machines, random forests, or deep learning architectures like convolutional neural networks can be employed for predicting various biological outcomes and modeling the behavior of biological samples under microfluidic applications [175, 176]. Following the acquisition of morphological image data from biosamples in MagLev microfluidic systems, machine learning techniques play a crucial role in various data processing tasks. Tracking the movement of particles or cells within the microfluidic channels, for instance, is essential for studying dynamic processes such as cell migration, proliferation, or drug response [177]. Machine learning algorithms, including particle tracking algorithms based on feature extraction or deep learning-based object detection methods, can accurately track individual particles or cells in real-time, enabling precise quantification and analysis of their behavior [178]. Machine learning algorithms can help identify patterns of variability within the data, allowing to distinguish between different cell types, states, or biomechanical features [179]. Segmentation and classification of biological structures within maglev microfluidic images are fundamental tasks for automated analysis and interpretation. Machine learning-based segmentation algorithms, such as U-Net, Mask R-CNN, or watershed transform coupled with neural networks, can accurately delineate individual cells, particles, or other relevant structures within images, facilitating subsequent quantitative analysis [180]. Moreover, machine learning algorithms can automate various aspects of experimental procedures, from sample loading and mixing to cell sorting and drug screening, reducing manual intervention and improving overall efficiency [176]. Reinforcement learning techniques, for instance, can adaptively control magnetic fields or fluid flow rates to optimize experimental conditions based on feedback from sensors or imaging systems, maximizing experimental throughput and reproducibility [181]. Real-time monitoring and analysis capabilities offered by machine learning algorithms are particularly beneficial in MagLev microfluidics, where rapid responses to dynamic changes are critical [182]. By continuously analyzing streaming data from sensors or imaging devices, machine learning models can detect anomalies, predict impending failures, or trigger adaptive interventions, ensuring the reliability and robustness of experimental setups [183].

Using MagLev technology in point-of-care (POC) devices involves analysis of the small volume of clinical samples within a paramagnetic medium [29]. The MagLev-enabled POC devices may accelerate the diagnostic decision-making process by not requiring lengthy or complex sample preparation or handling procedures [8]. These devices have the potential to revolutionize medical testing by enabling rapid and accurate assessments of various health parameters, ranging from detecting various biomarkers to monitoring disease progression and response to therapy [125, 184]. Moreover, these POC devices offer low-cost and easy imaging with optical microscopes, portable imaging systems for real-time observation and analysis [185].

Moreover, by integrating MagLev into telemedicine platforms, patients can have access to more advanced and real-time diagnostic capabilities from the comfort of their homes [86]. This innovation will be particularly valuable for chronic disease management, remote patient monitoring, and in situations where immediate access to healthcare expertise is vital [186, 187]. As telemedicine continues to evolve and expand, MagLev technology may play a pivotal role in ensuring high-quality healthcare services are accessible to individuals, regardless of their geographical location, contributing to improved healthcare outcomes and more efficient healthcare systems [188, 189].

5 Conclusion

MagLev technology is undergoing rapid development, marked by an exponential increase in novel applications and discoveries. Although the developed systems are generally non-flow systems, research has begun to focus on the dynamic applications under fluid flow. Current MagLev systems may be insufficient to achieve high resolutions that can detect very small density differences. Additionally, small-size particles cannot be levitated in a certain band due to Brownian motion in the MagLev system. To address these challenges, future research should prioritize the development of high-throughput analysis, the integration of hybrid systems with AI, and the advancement of compact and powerful magnets (Fig. 4). Having a significant capability for density-based separations and diagnostics, MagLev continues to take firm steps toward the future as an exciting technique that offers label-free analysis, rapid detection, simple operation, and an affordable platform. MagLev has the potential to not only redefine existing analytical standards but also create new pathways for scientific study and technological developments. The rate at which MagLev is developing emphasizes its significance as an innovative force, opening the way for a day when accessibility, efficiency, and precision will come together in ways that have never been seen before.

Fig. 4
figure 4

Envisioned advancements in MagLev technology. Integration of microfluidics, high-throughput analysis, and diverse biomarker profiling spanning from cellular to nucleic acid levels, alongside AI integration, are anticipated to enhance the capabilities of traditional MagLev systems

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