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can be created. In order to reduce the acquisition time, it is possible to create a static field with a “field‐free line” (FFL), but at the cost of resolution. The as‐obtained image represents the spatial distribution of SPIONs since the amplitude of the signal is proportional to the amount of magnetic material, usually expressed in Fe concentration. The foremost advantage is that the recorded signals do not depend on the aggregation state of MNPs, which in most biological applications cannot be neither monitored nor detected. This finding represents the huge advantage of MPI over MRI.

      One can consider that MPI is a development of Magnetic Nanoparticle Spectroscopy (MPS) (Wu et al. 2019). This very recent technique can be defined as a zero‐dimensional MPI scanner. Like in MPI, a sinusoidal magnetic field is applied to the SPIONs and their magnetization will be periodically driven in and out of saturation. The nonlinear SPIONs' magnetic response, containing unique spectral information, is recorded and separated into its spectral components. MPS has been actively explored as a portable, highly sensitive, cheap, in vitro, and easy‐to‐use bioassay testing kit. The MPS spectra (including harmonic amplitudes and phases) are unique for each type of SPIONs (Tu et al. 2014). In this manner, by conjugating the SPIONs with specific antibodies, it is possible to label different target analytes (even different cells) with specific types of SPIONs. This feature can be also used in MPI, as we will show later.

      2.2.3 MPI Cell Tracking

      The first major application of MPI was cell tracking (Bulte 2019). As mentioned above, the MPI's signal intensity linearly depends on MNPs' concentration (iron content for pure magnetite or maghemite) regardless of their state, their uniform distribution in a solution or their aggregation in clusters. This property is essential for biomedical applications because it is well known that in a biological environment the MNPs are either attached to cell surfaces or agglomerated in cytosol or endosomes after internalization by the cell. In the case of MRI, the aggregation state of the MNPs strongly influences the relaxation times, large clusters leading to a significant decrease of the observed transverse relaxation time.

      Usually, cell tracking is performed by loading the cells with MNPs. The distribution of these MNPs in the body will be monitored by MPI. It was reported that neural cells loaded with MNPs were detected in the rat brain. Moreover, the detection has been reconfirmed after several months from the first measurement (Zheng et al. 2016). Another important feature of MPI with respect to MRI is its higher sensitivity. The sensitivity depends mainly on the magnetic momentum of MNPs which in turn depends on the saturation magnetization and crystal volume. Janus‐like MNPs specially synthesized for MPI applications proved to be three times more effective as compared to Resovist and seven‐time more effective as compared to Feraheme (Song et al. 2018).

      While using MRI for tracking purposes, the cell's average iron content is 5–10 pg cell−1; in MPI, an entire field of view (FOV) in the case of a rat body measurements needs around 250 cells or 1 pg of iron. This higher sensitivity facilitates, at least from a theoretical point of view, the possibility to attain single cell tracking by using the MPI technique. For the above mentioned Janus MNPs, an in vivo sensitivity of 250 cells within the FOV of an entire mouse was reported (Song et al. 2018). This type of experiment cannot be visualized with MRI.

      Due to the EPR effect, systemic injection of the MNPs can lead to their accumulation within the tumor, allowing the tumor detection with a tumor‐to‐background ratio of 50 at the peak, six hours after the injection (Yu et al. 2017).

Schematic illustration of MPI-CT imaging of intravenously injected hMSCs, Resovist, and saline control, with representative coronal, sagittal, and axial slices shown from full 3D MPI datasets.

      Source: Reproduced with permission from Zheng et al. (2016).

      The versatility of the MPI technique allows the use of several classes of MNPs with different magnetic responses at different frequencies. In this manner, it is possible to separately collect multicollor MPI images for the different subclasses by assigning them different colors. This creates the possibility to concurrently study in vivo the interaction between the cells loaded as well as the MNP classes they are loaded with (Rahmer et al. 2017).

      Another important application of MPI is represented by MPI‐guided hyperthermia. The main advantage of using MPI in MH is the ability to check MNPs' distribution at the site of the tumor before applying the external magnetic fields. This is a real advantage, particularly when the MNPs are administered systemically (Tay et al. 2018). Moreover, it was observed that if the AC magnetic fields used for MPI, exhibiting ~20 kHz frequency, are applied alone no heating of the tissues occurs. However, if the AC frequency is increased to 340 kHz, heating of the tissue can be easily detected. At this point, the application of gradient magnetic fields will generate heat only in the FFP or FFL regions. As such, by positioning the FFP or FFL in the position of the tumor one might obtain heating only in the tumor area, without affecting neighboring healthy tissues.

      2.2.4 MNPs in Magnetic Hyperthermia

      The current standard of cancer care comprises the elimination of solid tumors by surgery followed by treatment with chemotherapeutic drugs (Bhattacharjee et al. 2010). However, it has been shown that most of anticancer drugs used in chemotherapy also target other healthy tissues in the body, causing major toxicity problems to vital organs. A typical example is the treatment with doxorubicin that provokes major heart problems as side effect (Minotti et al. 2004). Hyperthermia therapy or thermotherapy was considered a potentially useful alternative of chemotherapy. In this case, the body tissues are exposed to higher temperatures in order to damage or kill cancer cells by inducing cell apoptosis (Kerr et al. 1994). The concept of hyperthermia has been introduced in “clinical” practice many centuries ago by Greeks, Egyptians, Romans, and Indians. During the nineteenth century, it was observed that fever can cause tumor regression (Moyer and Delman 2008) and scientific studies were performed to treat cervical cancer by hyperthermia (Baronzio and Hager 2006).

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