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Logo of nihpaAbout Author manuscriptsSubmit a manuscriptHHS Public Access; Author Manuscript; Accepted for publication in peer reviewed journal;
Nanoscale. Author manuscript; available in PMC 2014 April 21.
Published in final edited form as:
PMCID: PMC3645980

Synthesis of ligand-functionalized water-soluble [18F]YF3 nanoparticles for PET imaging


We report a simple, efficient synthesis of novel 18F-labeled imaging agents based on YF3 nanoparticles. Targeting ligands and antitumor drug molecules can be introduced onto the YF3 nanoparticles in a one-pot synthesis. The 18F-labeling reaction proceeds in aqueous solutions at room temperature with excellent radiolabeling yields (> 80%) in very short time (5–10 min). 18F-labeled YF3 nanoparticles displayed high stability in mouse & human serum, and their application for mapping lymph nodes in live rats after local injection has also been demonstrated.

Rare-earth fluoride nanoparticles have attracted tremendous interest over the last few years because of their unique luminescence properties.14 Among all the studied rare-earth fluoride nanoparticles, yttrium trifluoride (YF3) nanoparticles have received much attention due to their potential applications as new laser materials and up-conversion imaging labels.57 Li et al. reported the preparation and up-conversion luminescence properties of fullerene-like and rice-like Ln3+-doped YF3 nanoparticles.8,9 Wang et al. studied the multicolor up-conversion emission in Yb3+/Tm3+, Yb3+/Ho3+ and Yb3+/Er3+ doped YF3 particles.10 Up-conversion YF3:Tm3+/Yb3+ nanocrystals were reported to emit from blue to magenta.11 These YF3 nanoparticles synthetic procedures often led to particles with irregular shapes, relatively large sizes or hydrophobic surface coatings. Nanoparticles with a smaller size, uniform shape, and good aqueous solubility are preferred for biolabeling and imaging applications.

On the other hand, optical imaging has limited tissue penetration in comparison to other imaging modalities such as positron emission tomography (PET). Biomolecules have been commonly radiolabeled for PET imaging studies of the activity of target receptors in living subjects.1216 Fluorine–18 (18F) is often used for PET imaging due to its ease in production in high quantities on a medical cyclotron and an ideal half–life of 110 min, but its introduction to the target molecule generally requires multiple synthetic steps often under harsh conditions and tedious purification processes.1719 Recently, the reaction between fluoride and rare-earth metal ions has been applied to label NaYF4 nanoparticles with 18F-.2022 These NaYF4 nanoparticles were made at high temperatures (320 °C) or in organic environments (in oleyl amine), and had to be converted into biocompatible nanoparticles through additional synthetic processes such as a ligand-exchange process of oleic acid with citrate. Generally, such multi-step strategies are associated with some intrinsic limitations, such as relatively high cost, and complicated preparation and post-treatment procedures. In this work, we describe a simple, efficient approach to synthesize 18F-labeled imaging agents based on YF3 nanoparticles (Scheme 1), the whole process including nanoparticles synthesis and 18F-labelling are performed in aqueous solution. Targeting ligands and even drug molecules were introduced to the nanoparticles in a one-pot synthesis. We also demonstrated their application for mapping lymph nodes in rats.

Scheme 1
Synthesis of [18F]YF3 nanoparticles.

Citric acid stabilized YF3 nanoparticles were synthesized by a modified coprecipitation procedure (ESI).23,24 The size and shape of synthesized nanoparticles are dependent on the ligand concentration, reaction time, and temperature. For example, when the concentration of citric acid decreased from 1 mmol to 0.5 mmol, the size range of Cit-YF3 nanoparticles increased from a range of 30–40 nm (Fig. 1A) to 30–80 nm (Fig. 1B). Increasing both reaction temperature and time led to larger Cit-YF3 nanoparticles; for example, by increasing the reaction temperature from 75 °C to 90 °C and the reaction time from 15 min to 60 min, the average size of Cit-YF3 nanoparticles increased from 30–40 nm to 50–70 nm (Fig. 1C).

Fig. 1
The transmission electron microscopy (TEM) images of theYF3 samples. (A) Cit-YF3: citric acid 1 mmol, 75 °C, 15 min, (B) Cit-YF3: citric acid 0.5 mmol, 75 °C, 15 min, (C) Cit-YF3: citric acid 1 mmol, 90 °C, 1 h, (D) the high resolution ...

Targeting ligands may be introduced during the synthesis. Folate was used together with citric acid as co-ligands to synthesize folate-conjugated YF3 nanoparticles (FA-YF3) for potentially targeting tumors over expressing folate receptors.2 TEM showed that the average diameter of FA-YF3 nanoparticles slightly decreased to 20–30 nm (Fig. 1E), and no obvious change in size was observed when the concentration of folate increased from 0.1 mmol to 0.2 mmol. High resolution TEM showed that the YF3 nanoparticles are composed of several smaller nanocrystals of ~ 5 nm (Fig. 1D&F). The energy-dispersive X-ray analysis (EDXA) pattern of FA-YF3 samples confirmed the presence of Y, F, O and N in the nanoparticles, with the additional Cu peak being readily attributed to the copper grid used (Fig. S1).

In addition to the targeting ligands, anticancer drug molecules such as doxorubicin (DOX) could be loaded onto the surface of FA-YF3 nanoparticles via electrostatic25 and coordination26 interactions to produce DOX-FA-YF3 (Scheme 1). After DOX conjugation, slight aggregation was observed from TEM images (Fig. S2). Dynamic Light Scattering (DLS) showed that lognormal size distribution of nanoparticles increased from 36 nm to 73 nm after DOX conjugation (Fig. S3). FA-YF3 nanoparticles exhibited an absorption peak at 281 nm pertaining to folate, and DOX-FA-YF3 nanoparticles exhibited two additional absorption peaks at 231 nm and 478 nm pertaining to DOX besides the folate absorption peak at 281 nm (Fig. 2A). These absorption peaks can be applied to determine the amount of folate and DOX loaded on the YF3 nanoparticles, which was about 4 ng/μg and 0.1 μg/μg, respectively (Fig. S4&5).

Fig. 2
(A) The UV-Vis absorbance spectra of the synthesized YF3 samples. (B) Cell viability values (%) estimated by MTT proliferation test versus incubation concentrations of nanoparticles and free DOX, respectively. MDA-MB-468 cells were cultured in the presence ...

In vitro cytotoxicity of YF3 nanoparticles was measured using the MTT assay against human breast cancer cell line MDA-MB-468 (Fig. 2B). Both Cit-YF3 and FA-YF3 showed low cytotoxicity: their cellular viability was estimated to be 73.8% and 79.4%, respectively, using the same concentration of 200 μg/mL nanoparticles. However, the viability of MDA-MB-468 cells incubated with DOX-FA-YF3 nanoparticles at the same concentration was estimated to be 20.3%, demonstrating that DOX was successfully loaded onto the YF3 nanoparticles and the loaded DOX remained cytotoxic. In comparison, free DOX exhibited a slightly higher toxicity, with 11.9% cellular viability at the same drug concentration.

18F-labeling was carried out by simply mixing [18F]KF solution with aqueous solutions of YF3 nanoparticles at room temperature followed by 5 to 10 min incubation, and free 18F was easily removed by centrifugation. YF3 nanoparticles with different surface ligands, including citric acid (Cit-YF3), folate (FA-YF3), DOX (DOX-FA-YF3) and PEG linkage (PEG2000-Cit-YF3) were 18F-labeled using this method. Excellent radiolabeling yields were observed, generally in the range of 80–95% (decay corrected to end of bombardment) (Fig. 3A). The concentration of nanoparticles and increased reaction time did not lead to significant increases in the radiolabeling yields. For example, the 18F-labeling yield for Cit-YF3 nanoparticles was 96%, 95%, and 84% with just 5 min reaction time at the concentration of 1, 0.5, and 0.2 mg/mL, respectively, and no obvious change in the radiolabeling yield was observed when the incubation time increased to 20 min (Fig. S6). We examined the serum stability of 18F-labeled YF3 nanoparticles by measuring dissociated [18F] fluoride in the supernatant. Only 6% of [18F]fluoride was released from Cit-[18F]YF3 nanoparticles during first 30 min incubation in mouse and human serum, which might be due to the loss of fluoride non-specifically absorbed on the nanoparticles, and nearly no further [18F] fluoride dissociation was observed in prolonged incubations of 2–6 h (Fig. S7).

Fig. 3
(A) 18F labeling yield of synthesized YF3 nanoparticles. (B) Comparison of uptakes of FA-[18F]YF3 nanoparticles in PYMT and MDA-MB-468 cell lines.

To evaluate the targeting effect of folate-conjugated nanoparticles, the uptakes of FA-[18F]YF3 nanoparticles in PyMT (polyoma middle T oncoprotein mouse breast cancer, folate receptor positive)27 and MDA-MB-468 cell lines (folate receptor negative)28 were investigated. FA-[18F]YF3 showed 2–3 fold higher cellular uptake efficiency in PyMT cells compared with that in MDA-MB-468 cells at 45 min, 1 h, and 2 h, respectively (Fig. 3B).

Finally labeled [18F]YF3 nanoparticles were applied to map lymph nodes in vivo. The lymphatic system plays a crucial role in immune responses to foreign antigens and tumors, and in tumor metastasis in human and rodent models.29 Two different sizes of Cit-[18F]YF3 nanoparticles, one in the range of 30–40 nm in diameter and the other at 50–70 nm, were evaluated by intradermal injections into the right and left front footpads of rats. Both axillary lymph nodes could be easily visualized non-invasively by microPET/CT imaging at 15 min post injection (Fig. 4 and Fig. S8). The axillary lymph node uptake of smaller nanoparticles was 6.7 times greater than that of larger nanoparticles. Similarly, an 8.4 times higher uptake of smaller nanoparticles in the popliteal lymph nodes was observed when these Cit-[18F]YF3 nanoparticles were injected into the rear footpads of rats (Fig. S9). The size of nanoparticles exhibits a strong effect on the migration time during lymph node mapping, and nanoparticles with a hydrodynamic diameter of 10–50 nm were reported to exhibit rapid uptake into the lymphatic system and considered optimal for lymph node mapping30,31 Our study further confirmed this result.

Fig. 4
Imaging lymph nodes in rats with [18F]YF3 nanoparticles after 15 min injection at front foot pads (indicated by dotted arrows); axillary lymph nodes are indicated by solid arrows.

In conclusion, we have reported a simple, efficient procedure to synthesize water dispersible 18F-labeled YF3 nanoparticles, which avoided high temperatures, high pressures, and organic solvents. Targeting and antitumor drug molecules can be introduced onto the nanoparticles for 18F-labeling. The labeling reaction proceeds in aqueous solutions at room temperature with excellent radiolabeling yields (> 80%) in very short time (5–10 min). Labeled YF3 nanoparticles showed high serum stability and were able to map lymph nodes in live rats after local injection. This simple, efficient 18F-labeling strategy should enable the use of rare-earth nanoparticles for both PET and optical imaging in vivo.

Supplementary Material



This work was supported by the NIH National Cancer Institute (NCI) grants R01CA135294, R21CA138353A2, the Stanford University National Cancer Institute (NCI) CCNE-T grant (U54CA119367) and ICMIC (P50CA114747). We acknowledge the use of the SCi3 Core Facility.


Electronic Supplementary Information (ESI) available: [details of the experiment section as well as EDXA, TEM, DLS, UV-Vis spectra, 18F labeling yield, serum stability, biodistribution, and lymph node images]. See DOI: 10.1039/b000000x/


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