Bi 2 O 3 boosts brightness, biocompatibility and stability of Mn-doped Ba 3 (VO 4 ) 2 as NIR-II contrast agent † Materials Chemistry B

Deep-tissue fluorescence imaging remains a major challenge as there is limited availability of bright biocompatible materials with high photo- and chemical stability. Contrast agents with emission wavelengths above 1000 nm are most favorable for deep tissue imaging, oﬀering deeper penetration and less scattering than those operating at shorter wavelengths. Organic fluorophores suﬀer from low stability while inorganic nanomaterials ( e.g. quantum dots) are based typically on heavy metals raising toxicity concerns. Here, we report scalable flame aerosol synthesis of water-dispersible Ba 3 (VO 4 ) 2 nanoparticles doped with Mn 5+ which exhibit a narrow emission band at 1180 nm upon near-infrared excitation. Their co-synthesis with Bi 2 O 3 results in even higher absorption and ten-fold increased emission intensity. The addition of Bi 2 O 3 also improved both chemical stability and cytocompatibility by an order of magnitude enabling imaging deep within tissue. Taken together, these bright particles oﬀer excellent photo-, chemical and colloidal stability in various media with cytocompatibility to HeLa cells superior to existing commercial contrast agents.


Introduction
Luminescent nanoparticles for bioimaging are most attractive for non-invasive acquisition of anatomic and functional information at high spatio-temporal resolution. Deep-tissue luminescence imaging is accomplished by probes with emission wavelengths either in the first 1 (650-900 nm) or second 2 (1000-1350 nm) near-infrared (NIR) windows. At these wavelengths, light absorption 3 and scattering 4 are reduced compared to visible light wavelengths enabling improved imaging depth. Moreover, the autofluorescence from endogenous fluorescent molecules in biological tissues is weaker. 5 Tsukasaki et al. 6 demonstrated how the combination of these properties favorably influences the signal to background ratio, increasing it from 1.3 (emission at 720 nm) up to 7.3 (emission at 1300 nm) in an in vivo imaging setting.
An ideal fluorescent probe for deep-tissue imaging should possess high quantum yield (QY), as well as photostability, biocompatibility and chemical stability. 7 Furthermore, a large Stokes shift and narrow emission lines hinder reabsorption and emission cross-talk with other emitters. 7 Fluorescent contrast agents in the second NIR window include organic molecules (dyes 8 and conjugated polymers 9 ) typically suffering from limited photostability, 10 and single-walled carbon nanotubes 11 (SWCNT) with broad size distribution 12 (200-500 nm length) and potentially cell-damaging needle-like structure. 7 Furthermore, there are semiconductor quantum dots often consisting of toxic heavy metals (e.g. InAs, 13 PbS 14 ), and lanthanide-doped materials limited by the small absorption cross-section, 15 which however can be improved by sensitizing via organic dyes 16 or co-doping. 16,17 An almost unexplored group of materials for NIR-II bioimaging are transition metal doped materials, despite several studies reporting emission above 1000 nm in relatively large crystals (micrometer to millimeter size): divalent (Ni, 18 Co, V, 19 and Ti 20 ), trivalent (V 21 ), tetravalent (Cr, 22 V), pentavalent (Cr, 23 Mn 24 ), and hexavalent (Mn, 25 Fe 26 ) transition metal ions exhibit attractive properties. Especially Mn 5+ as an active ion doped into an otherwise NIR-inactive Ba 3 (VO 4 ) 2 matrix is most promising due to its broad excitation band spanning the entire first biological window and its strong and narrow emission peak around 1180 nm. 27 Only recently Zhang et al. 28 have reported synthesis of submicron particles (mean diameter B300 nm) of Ba 3 (VO 4 ) 2 and Ba 3 (PO 4 ) 2 doped with Mn 5+ leading to NIR emission, however, the cytocompatibility of these materials remains unexplored, and particle sizes exceed 300 nm, making them unsuitable for intravascular applications such as passive tumor targeting. 29 Specifically, particle sizes around 120 nm have been found suitable for tumor imaging, 30 as well as metastatic tiny tumor detection and tumor vessel visualization. 31 We recently made smaller Ba 3 (PO 4 ) 2 :Mn 5+ particles (d o 100 nm) with the potential for simultaneous deep-tissue imaging and nanothermometry. 32 Despite encouraging results, these particles show partial dissolution (10% in 24 hours) in physiological media such as PBS (phosphate buffered saline), and their cellular internalization has not yet been studied.
Here, we present Ba 3 (VO 4 ) 2 :Mn 5+ nanoparticles with sizes below 100 nm for the first time made by sterile flame aerosol technology for its close control of product particle characteristics. 33 This technology is most effective in overcoming the typical clinical translation barriers, 34 such as scalability and reproducibility. 35 Most importantly, we demonstrate how co-synthesis with Bi 2 O 3 improves drastically both brightness and stability of Ba 3 (VO 4 ) 2 :Mn 5+ . The resulting particles show high cytocompatibility (HeLa cells) due to reduced ion leaching. By assessing structure-function relationships, we provide important new insights into the mostly unexplored group of Mn 5+ -doped materials and their suitability for bioimaging.

Results and discussion
Optical properties of Ba 3 (VO 4 ) 2 :Mn 5+ and influence of Bi 2 O 3 to brightness Flame spray pyrolysis (FSP) has been employed for synthesis of Ba 3 (VO 4 ) 2 particles doped with Mn 5+ at typical production rates of 10 g h À1 , which could be scaled up to kg h À1 as shown with a variety of materials. [36][37][38] Following extensive optimization in terms of doping concentration and particle size ( Fig. S1-S11, ESI †) Ba 3 (VO 4 ) 2 doped with 0.5 at% Mn 5+ , Ba 3 (V 0.995 Mn 0.005 O 4 ) 2 , annealed for 2 h at 600 1C (abbreviated from now on as BaVOMn) was found to be most suitable with a quantum yield (QY) of 10.6 AE 2.1% as powder and 4.6 AE 1.1% in aqueous suspension. The QY was directly measured by an absolute method using an integrating sphere (Fig. S12, ESI †) which avoids the need for reliable standard materials that are scarce in the NIR region. For example, the only available reference-dye for relative QY measurements in the NIR (IR26) has been shown to have a QY that varies by an order of magnitude between different reports. 39 Fig . 1a shows the excitation (dotted line) and emission spectra (solid line) for BaVOMn. The excitation spectrum features a narrow peak in the UV and a broad one spanning the entire first NIR biological window. There are three spin-allowed transitions from the 3 A 2 ground state that dominate the excitation and absorption spectra at 341, 654 and 888 nm, as well as a weaker spin-forbidden transition at 777 nm and a charge transfer band (CT) in the UV from the host (details in Fig. S7 and S8, ESI †). In contrast to lanthanide-based materials with narrow excitation peaks, 40 the broad absorption band in NIR allows more freedom in the choice of excitation source. For example, 750 or 808 nm lasers can be employed in contrast to 980 nm (a typical excitation wavelength for Yb-based emitters 41 ) to avoid overheating due to the strong optical absorption of water. 42 The emission peak is centered around 1181 nm and is assigned to the 1 E -3 A 2 transition. 24 The weak side emissions correspond to vibronic sidebands. 43 These emissions lie within the second biological window, therefore allowing deeper tissue penetration compared to the first biological window. The fullwidth at half-maximum (FWHM) of the main emission line is only 10 nm and thus an order of magnitude lower than for QDs (around 200 nm 14 ). This can be beneficial in avoiding unwanted cross-talk 7 between several emitters in a multicolor imaging setting. It is noteworthy that the observed absorption and excitation/emission spectra unambiguously indicate the presence of Mn 5+ . This is further supported by the blue coloration of the powders (Fig. S6, ESI †), as well as Raman spectral fingerprints characteristic for Mn 5+ (Fig. S9, ESI †).
To further increase the emission intensity (i.e. brightness of the particles at identical particle concentration), co-synthesis of BaVOMn with bismuth oxide has been explored, as the latter increases the emission intensity of several phosphors. 44,45 A Bi-precursor was added to the precursor solution of BaVOMn prior to its FSP yielding BaVOMn -y wt% Bi 2 O 3 with y = 0 to 108.9 wt%, relative to the BaVOMn mass, where y = 108.9 wt% corresponds to a Ba to Bi molar ratio of 1 : 1. The fluorescence emission intensities of aqueous dispersions of these particles at constant concentration (0.5 mg ml À1 ) are shown in Fig. 1b, normalized to that of BaVOMn alone. The presence of Bi 2 O 3 drastically improved the emission intensity of these already bright particles. The highest brightness was observed for 54.4 wt% Bi 2 O 3 that exhibited an almost tenfold higher intensity than pure BaVOMn. The shape of the emission spectra was not affected by the addition of Bi 2 O 3 . Note that neither Ba 3 (VO 4 ) 2 (without Mn) -1 wt% Bi 2 O 3 nor Bi 2 O 3 : 1% Mn exhibited any luminescence. The brightest sample with 54.4 wt% Bi 2 O 3 was abbreviated with BaVOMn-BiO.
With increasing Bi 2 O 3 content, y, the absorption (triangles) at the excitation wavelength of 750 nm was increased (Fig. 1c). The stabilization of Mn 5+ has been shown to be difficult, depending e.g. on small changes in host stoichiometry 32 or annealing temperature, which was also observed here in samples without Bi 2 O 3 (Fig. S6c, ESI †). The absorption spectra (Fig. S13, ESI †) indicate that the addition of Bi 2 O 3 favors stabilization of Mn 5+ in the host material, thereby increasing the absorption peak in the near-infrared. However, this stabilization effect appears to level off for higher Bi 2 O 3 contents, leading to the observed peak at 54.4 wt% Bi 2 O 3 followed by a decrease due to the addition of inactive mass.
Next, the efficiency of these particles is studied in detail with their QY, for different Bi 2 O 3 content, as shown in Fig. 1c (circles) and compared with other reported emitters in this spectral region in Table S1 (ESI †). The QY increases with Bi 2 O 3 content and peaks at 54.4 wt%, similar to absorption. Compared to our previously developed 32 Ba 3 (PO 4 ) 2 :Mn 5+ , the NIR absorption of BaVOMn-BiO is more than doubled at comparable QY.

Physicochemical characterization and colloidal stability
Motivated by the improved optical properties of BaVOMn-BiO over BaVOMn, the physicochemical properties of BaVOMn-BiO were studied in detail. The addition of Bi 2 O 3 hardly affected the crystal size but increased the primary size (Fig. S14a, ESI †). Therefore, the annealing conditions were investigated ( Fig. S14-S16, ESI †) for the most promising sample according to Fig. 1b and c, BaVOMn-BiO, to find those at which the particles show phase purity yet minor amount of aggregation. Such conditions were found at 550 1C for 30 minutes, with the crystal (d XRD = 58 nm) and primary particle (d BET = 56 nm) sizes close to each other. Due to the smaller crystal size, also the QY of this powder decreased to 8.1 AE 0.5% (2.7 AE 0.3% in water), yet represents a compromise between brightness and suitable particle size (i.e. less than 100 nm). Fig. 2a shows a high-angle annular darkfield scanning transmission electron microscopy (HAADF-STEM) image of BaVOMn-BiO nanoparticles. The Z-contrast reveals inhomogeneities in the particles, evidenced by differences in brightness. The small bright dots (o5 nm) are metallic Bi-domains that formed during imaging (Fig. S17, ESI †). In contrast, the larger inhomogeneities appeared during the thermal treatment and could not be observed in the as-prepared samples (Fig. S18, ESI †). The same particles were analyzed with energy dispersive X-ray spectroscopy (EDX) elemental mapping. Fig. 2b shows overlay of V, Ba and Bi maps (single element maps are shown in Fig. S19, ESI †). The Ba is colocated with V, while Bi is seggregated. Two areas of distinctly high (area 1) and low (area 2) Bi concentrations have been highlighted in Fig. 2b and the corresponding EDX-spectra are shown in Fig. 2c and d, respectively. They clearly confirm the strongly varying ratio of Ba to Bi, however, even in these extreme regions always both elements can be detected. The segregation of Bi-rich areas could potentially be explained by the lower melting point of Bi 2 O 3 (825 1C) 46 than Ba 3 (VO 4 ) 2 (1600 1C) 47 and thus its higher mobility. The Mn distribution appears to be rather uniform (Fig. S19e, ESI †). Fig. 3a shows the XRD patterns of both powders annealed at 600 1C. For BaVOMn, all peaks can be attributed to rhombohedral Ba 3 (VO 4 ) 2 (circles), while additional peaks corresponding to tetragonal Bi 2 O 3 (triangles) can be observed in BaVOMn-BiO, in line with the observed presence of Bi rich areas in the elemental mapping (Fig. 2b). The XRD analysis of varying Bi 2 O 3 concentration (Fig. S20, ESI †) reveals Bi 2 O 3 peaks for y = 5.4 wt% that increase in intensity with increasing Bi content. For lower y either the signal is below detection limit or Bi 3+ substitutes the 8-coordinated Ba 2+ (ionic radii: Ba 2+ = 142 pm, Bi 3+ = 117 pm). Analysis of the unit cell volume (Fig. S21, ESI †) showed a slight increase from 0 to 10.9 wt% Bi 2 O 3 leveling off above that, indicating potential incorporation of Bi. This is supported by the observed Bi-presence from electron microscopy even in barium rich areas ( Fig. 2b and d).
The amount of Bi in the nanoparticles was close to the nominal one, as determined by ICP-OES (Fig. S22, ESI †). Fig. 3b shows an exemplary TEM image and the corresponding size distribution with mean diameter and geometric standard deviation of 62 nm and 1.29, which is in good agreement with measurements by XRD (d XRD = 58 nm) and N 2 adsorption (d BET = 56 nm). Their shape is slightly elongated, originating from sintering and aggregation of primary particles. In fact, elongated particles (i.e. discoidal and cylindrical) were most attractive for targeting tumors as they had accumulated more than spherical or quasi-spherical ones to organs (liver, spleen and lungs) of tumor-bearing mice. 48 These particles were coated with human serum albumin (HSA) to improve their colloidal stability 49 due to the extra electrostatic repulsion provided by the proteins, 50 as well as the potential benefit of increased blood circulation times due to lower non-specific interaction. 51 The successful functionalization was confirmed with FTIR and TGA (shown in Fig. S23 and S24, ESI †). Following differential centrifugation (Fig. S25a, ESI †) to remove the undesirable fraction (80 wt%) of largest aggregates formed during annealing (yield B20 wt%), hydrodynamic particle size distributions centered between 60 and 200 nm with polydispersity index around 0.1 have been obtained (Fig. S25b and c, ESI †), shown exemplarily in Fig. 3c in water. The successful separation of various size fractions by centrifugation and the removal of larger aggregates has been confirmed by electron microscopy at two magnifications ( Fig. S26a-d, ESI †). The resulting aggregate size distributions of the largest Feret diameters determined by microscopy compared well with DLS measurements ( Fig. S26e and f, ESI †), given the different measurement principles. 52 As mentioned above, such hydrodynamic sizes are suitable for blood flow mapping (e.g. d average = 330 nm by Bruns et al. 13 ) as well as metastatic tumor detection, i.e. d = 30-450 nm by Gao et al. 30 and d = 90-140 nm by Li et al. 31 These distributions were monitored for up to 30 days in water, 0.154 M NaCl as well as RPMI cell culture medium (Fig. 3d). The hydrodynamic mode size remains constant in all media, without significant changes in size distribution (Fig. 3c: as prepared, t = 0, and after 30 days), indicating high colloidal stability (Fig. S27, ESI †). Furthermore, the evolution of their luminescence intensity in different media (water, PBS, RPMI medium) over 14 days (Fig. S28a, ESI †) showed no sign of deterioration of performance. Additionally, that intensity remained constant over pH = 5-10, for at least 14 days (Fig. S28b, ESI †).

In vitro characterization and biocompatibility
The cytocompatibility of nanoparticles (concentration: 0-1000 mg ml À1 ) was assessed in vitro by incubating them with  Fig. 4a, BaVOMn particles (blue bars) cause a reduction in cell viability with the LD50 around 55 mg ml À1 . Similarly, also commercial PbS-CdS QDs functionalized for bio-applications (obtained from NanoOptical Materials) were tested as a benchmark, since these have been studied extensively for fluorescence microscopy including deeptissue imaging. 14 The PbS-CdS QDs (orange bars) showed a reduction in cell viability reaching below 75% at 100 mg ml À1 . Higher QD concentrations could not be determined, as the concentration of supplied colloids was not high enough. Remarkably, the BaVOMn-BiO particles (red columns) exhibited superior cell compatibility. Even if corrected for the lower Ba 3 (VO 4 ) 2 content due to the Bi 2 O 3 mass contribution, the nanoparticles are still better tolerated than pure BaVOMn, notably at comparable sizes (see Table S3, ESI †). The HeLa cell cytocompatibility of the BaVOMn-BiO particles is comparable to that of less bright Ba 3 (PO 4 ) 2 :Mn 5+ nanoparticlesd (up to 1000 mg ml À1 ). 32 To gain a deeper understanding, also the cytotoxicity of water soluble BaCl 2 (green bars) and Na 3 VO 4 (pink bars) was tested to investigate the toxicity of barium and orthovanadate ions. The BaCl 2 hardly affected the viability of HeLa cells up to  250 mg ml À1 with significant impact only at the two highest concentrations (500 and 1000 mg ml À1 ). In contrast, even the lowest dose of Na 3 VO 4 led to reduction in cell viability down to 55%, resulting in an LD50 of 30 mg ml À1 . These results are in line with reported values for L929 fibroblasts, where the LD50 for vanadium salt was more than an order of magnitude lower than for BaCl 2 . 53 To assess whether there is a difference in the amount of released ions from BaVOMn and BaVOMn-BiO nanoparticles that would explain the distinctly different cytotoxicity, leached ions were quantified in three relevant media (water, PBS and cell culture medium RPMI) by ICP-OES (Fig. 4b).

HeLa cells for 24 hours. As shown in
Remarkably, the addition of Bi 2 O 3 increases the stability for BaVOMn-BiO (filled columns) with leached ion concentration almost an order of magnitude lower than pure BaVOMn (open columns), in all three media. The highest fraction dissolved was in PBS with 34.6% Ba and 33.7% V in BaVOMn. In contrast, there is a distinct reduction in ion release for BaVOMn-BiO, where in PBS only 3.2% of Ba and 1.9% V were dissolved. Since the HSA-coating density determined by TGA (Fig. S24, ESI †) was similar for BaVOMn (2.4 mg HSA m À2 ) and BaVOMn-BiO (1.5 mg HSA m À2 ), this was unlikely to explain such large differences in cytotoxicity and leaching behavior.

À1
). These values are still orders of magnitude lower than the median lethal dose (LD50) in mice of 19.2 mg kg À1 for barium 54 and 18.4 mg kg À1 for vanadium 55 after intravenous injection. Therefore, it can indeed be concluded that the increased cytocompabitility of BaVOMn-BiO is attributed to the reduced amount of leached ions compared to BaVOMn. The Bi 2 O 3 appears to stabilize the nanoparticles and make them less prone to dissolution and release of Ba and V ions, thus making them very promising for in vivo applications. This could be caused by the occupation of vulnerable defect sites by Bi 2 O 3 , similar to black phosphorus/ Bi 2 O 3 heterostructures. 56 Furthermore, since Bi 2 O 3 nanoparticles functionalized with HSA have exhibited excellent biocompatibility in vivo, 57 this could further explain the observed improvement in cytocompatibility.
Bright particle emission within an intracellular environment was confirmed by fluorescence microscopy, where the interaction of these particles incubated with HeLa cells was further studied. Fig. 4c shows the overlay image of three different fluorescent signals: The cytoskeleton was stained with the dye Alexa 488 Phalloidin (l em = 530 nm) shown in green, the cell nucleus was stained with the blue-emitting dye DAPI (l em = 455 nm), and the emission of the BaVOMn-BiO particles at 1180 nm is shown in red. The nanoparticles accumulate in the perinuclear region, indicating that the nanoparticles readily enter the cell. Also, some extracellular particles are observed that are typical for in vitro culture settings where particle sedimentation is a dominant mechanism for particle-cell contact.

Benchmarking with commercial fluorescent NIR-II agents
To best assess the performance of BaVOMn-BiO, it is compared to commercially available materials for fluorescence bioimaging in the NIR-II, specifically to the organic dye indocyanine green (ICG) and PbS-CdS QDs coated with diol groups on the surface. Fig. 5a shows the emission spectra of aqueous suspensions of all three samples at equal mass concentrations (0.1 g L À1 ) under 750 nm excitation. This wavelength is not at the optimum for any of the three emitters (excitation spectra shown in Fig. S31, ESI †) but can excite all of them while still lying in the first NIR window. The QDs (green broken line) have a strong emission between 1000 to 1350 nm. The emission spectrum of these commercial NIR-II Pbs-CdS QDs is rather broad and unattractive due to the observed shoulder, and consistent to literature 14 (FWHM of B200 nm, their 14 Fig. 2a  and b). The BaVOMn-BiO nanoparticles (blue solid line) show an almost equal peak intensity, while the ICG emission (red dot-broken line) peaks in the first NIR window and has only a weak emission tail in the NIR-II region. Still, it has been demonstrated that using this tail leads to better contrast than using the emission peak in the NIR-I region for bioimaging. 58 Besides brightness, also stability is crucial for fluorescent contrast agents. 59 The photostability (at peak wavelength) for one hour under continuous excitation (750 nm, 2.9 W cm À2 ) of these three materials is shown in Fig. 5b. While the present BaVOMn-BiO shows stable emission even up to 12 hours (Fig. S32, ESI †), both ICG and the QDs drastically decrease their emission intensity within the first 20 minutes. For ICG, this is attributed to photobleaching. 60 Similar behavior has been observed by Zhao et al. 61 for these QDs (their 61 Fig. 9a) that was attributed to quenching at unpassivated surface sites. Since no blue-shift of the emission peak was observed, changes in size of the QDs were ruled out.
Finally, the potential of these particles for deep-tissue imaging was studied ex vivo using chicken skeletal muscle tissue and a defined structure simulating a small blood vessel. Fig. 5c shows the fluorescence images of glass capillaries 62 (diameter = 1.5 mm) filled with fluorescent contrast agents (BaVOMn-BiO, Pbs-CdS QDs and ICG) under layers of chicken tissue with increasing thickness (0-10 mm).
The contrast agents were excited with a 750 nm laser (37.4 mW cm À2 , one order of magnitude below the exposure limit 63 ) and images were acquired with a NIR-camera equipped with an 850 nm long-pass filter, where the exposure time was adjusted for each image close to saturation to maximize signal intensity.
The apparent width of the fluorescence image coming from the glass capillary without tissue (Fig. 5c at 0 mm tissue thickness) is clearly defined and starts broadening with increasing tissue thickness due to scattering. The latter was quantified by determining the FWHM (Fig. 5d). For both QDs and present nanoparticles, the FWHM increases from 1.3 mm up to B8 mm for the maximum tissue thickness of 10 mm, which is expected due to the similar spectral operating range, in agreement with phantom tissue experiments. 12 In contrast, the FWHM of ICG is much more affected by tissue scattering and reaches a maximum of 19.4 mm at 10 mm depth. This demonstrates the advantage of operating within the NIR-II window, in contrast to the NIR-I, where the main emission peak of ICG lies. Furthermore, also smaller features could potentially be detected with additional optics resulting in smaller field of view and reaching sub-10 mm spatial resolutions. 64 As an additional measure of image quality, the signal-tobackground ratio (SBR) was quantified in Fig. 5e, using images acquired with laser excitation but without fluorescent contrast agents as background. No significant difference between the present nanoparticles and QDs can be observed, while ICG shows a much smaller ratio. It is noteworthy that even at 10 mm of tissue thickness, the signal of the BaVOMn-BiO is 40 times higher than the background. According to the Rose criterion, 65 a minimum SBR of 5 is required to distinguish image features with 100% certainty. Thus, even weaker or lower lying signals down to B3 cm 32 can still be detected reliably.

Conclusions
Sub-100 nm Mn 5+ -doped Ba 3 (VO 4 ) 2 nanoparticles with bright emission in the NIR-II and high quantum yield were produced by scalable and sterile flame aerosol technology, thereby fulfilling key requirements for bioimaging. Co-synthesis with Bi 2 O 3 enhances their brightness up to 10-fold and drastically improves their chemical stability and cytocompatibility to HeLa Comparison of present BaVOMn-BiO to commercial fluorescent agents with emission in the NIR-II at equal mass concentration (0.1 g L À1 ) dispersed in water in terms of (a) emission intensity, and (b) photostability under constant illumination (750 nm, 2.9 W cm À2 ) at their respective peak intensity wavelength. (c) Ex vivo bioimaging: Glass capillaries filled with fluorescent contrast agents were placed under chicken skeletal muscle tissue of varying thickness, excited with a 750 nm laser and imaged. Extracted full-width at half-maximum (FWHM) (d) and signal to background ratios (e). Both BaVOMn-BiO and Pbs-CdS QDs show comparable performance. cells, thus representing an attractive material for bioimaging. The aforementioned properties make BaVOMn-BiO nanoparticles a most promising NIR-II contrast agent for deep tissue imaging as it surpasses traditional fluorophores and quantum dots in photostability while exhibiting better biocompatibility than quantum dots at similar brightness and imaging capabilities. While ex vivo experiments are suited better to systematically study the penetration depth and spatial resolution of novel contrast agents due to their homogeneity, further in vivo experiments are the next step towards clinical applications to fully address and unfold the performance of these BaVOMn-BiO particles.

Conflicts of interest
There are no conflicts to declare.