Lanthanide-based nanoparticles as vascular contrast agents for micro-computed tomography

Joy Dunmore-Buyze (Robarts Research Institute)
David W. Holdsworth (Robarts Research Institute)
Maria Drangova (Robarts Research Institute)
Elizabeth R. Gillies (Western University)


Vascular development is extensively studied in small animals with the goal of increasing our understanding of this process and applying what is learned to humans. Micro-computed tomography (micro-CT) is widely used to obtain high-resolution and non-destructive 3D images of small animal vasculature. To distinguish the vasculature from other soft tissue, long-circulating agents (i.e. > 10 minutes) containing a high contrast element loading (i.e. 100 mg/mL) are required. This can be achieved by using polymer-encapsulated nanoparticles exceeding 10 nm in diameter (1). Although nanoparticle-based agents exist for micro-CT, they are predominantly based on iodine, which has a low atomic number. Higher contrast can be achieved by using elements with higher atomic numbers, such as lanthanides (e.g. Er), particularly at higher CT energies. While lanthanide-based contrast agents are used clinically and in pre-clinical research, they are either composed of small molecules (< 1 nm) that exit the bloodstream of small animals within seconds, or do not contain sufficient quantities of lanthanide. Therefore, the objective of this work was to develop polymer-encapsulated Er nanoparticles exceeding 10 nm in size as a vascular contrast agent in micro-CT. We hypothesized that the synthesized agent will circulate in the blood pool of small animals for tens of minutes, while providing vascular contrast when imaged by micro-CT.

Materials and Methods

Contrast agent preparation and characterization- A series of polymers were studied to identify a formulation that remained stable in a mouse blood mimic and had relatively higher Er content than others. This polymer – which was a diblock copolymer containing poly(ethylene glycol) and poly(L-lactide) – was used to prepare encapsulations containing synthesized NaErF4 nanoparticles (ErNP) (2) by nanoprecipitation (3). The resulting suspension was filtered, freeze-dried and stored at room temperature. The ErNP were characterized using dynamic light scattering (DLS) and transmission electron microscopy (TEM). The dried formulation was dissolved in normal saline immediately before use at an Er concentration of 100 mg/mL.

In vivo application of the contrast agent- Using 0.2 mL of the contrast agent, male C57BL/6 mice (25-30 g) were injected subcutaneously (n=2) and intravenously (n=3) via tail vein catheterization. The biodistribution of the contrast agent was observed by micro-CT. The animals that were injected subcutaneously were sacrificed for gross examination of subcutaneous tissue.

Micro-CT imaging and analysis- Micro-CT images were obtained using the GE Locus Ultra (London, ON), where 1000 views (16 ms per view) were acquired at 80 kVp, 55 mA over 360° and reconstructed using a cone-beam reconstruction algorithm to a voxel size of 150 × 150 × 150 µm. Images were analyzed using MicroView (Parallax Innovations, London, ON) and CT contrast was reported in Hounsfield Units (HU).


Physical characterization- The polymer assemblies containing ErNP had an average hydrodynamic diameter of 171 ± 3 nm with low size dispersity as measured by DLS, and TEM results showed good agreement with the DLS-measured diameters (Figure 1). The optimized formulation was also confirmed to contain 100 mg/mL of Er.

In vivo characterization- After subcutaneous administration, the ErNP remained localized in the injection site for up to a week (Figure 2). No signs of irritation or necrosis were observed in the subcutaneous tissue upon gross examination. In the blood pool of the animals that were intravenously injected, contrast enhancements of over 250 HU were observed for up to an hour (Figure 3).

Discussion and Conclusion

Polymer-encapsulated Er nanoparticles that were larger than 10 nm in size and could encapsulate 100 mg/mL of Er were successfully synthesized. The contrast agent did not irritate subcutaneous mouse tissue for up to a week, and remained stable and inert in vivo. In the blood pool, contrast enhancement was observed for up to an hour; this well exceeds in vivo micro-CT requirements. This work represents the development of the first nanoparticle-based contrast agent that contains 100 mg/mL of Er and is targeted for in vivo micro-imaging.

Figure 1. A TEM image and the DLS-measured hydrodynamic volume size distribution of the contrast agent.

Figure 2. Dermal (left) and subcutaneous (right) tissue of a mouse that was injected with the agent. The arrows indicate the injection site and contrast agent localization.

Figure 3. Time-course micro-CT images of a mouse that was intravenously injected with the contrast agent. The blood pool in the heart - which was indistinct in the pre-contrast image - appears much brighter after injection. Contrast enhancement was observed for up to 60 minutes in the blood pool.


1. Choi CH, Zuckerman JE, Webster P, Davis ME. Targeting kidney mesangium by nanoparticles of defined size. Proc Natl Acad Sci USA. 2011;108(16):6656-61.

2. Zhao G, Tong L, Cao P, Nitz M, Winnik MA. Functional PEG-PAMAM-tetraphosphonate capped NaLnF(4) nanoparticles and their colloidal stability in phosphate buffer. Langmuir. 2014;30(23):6980-9.

3. Prashant C, Dipak M, Yang CT, Chuang KH, Jun D, Feng SS. Superparamagnetic iron oxide--loaded poly(lactic acid)-D-alpha-tocopherol polyethylene glycol 1000 succinate copolymer nanoparticles as MRI contrast agent. Biomaterials. 2010;31(21):5588-97.

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