A versatile platform for bioimaging based on colominic acid-decorated upconversion nanoparticles.

Lanthanide-doped upconversion nanoparticles (UCNPs) are promising bioimaging agents that emit light under near infra-red excitation, capable of penetrating deep in biotissues with a high signal-to-noise ratio. Their successful implementation is principally associated with surface functionalization. Here, we report on UCNP surface modification with highly hydrophilic, endogenous, non-toxic, non-immunogenic colominic acid, conferring "stealth" properties. We proposed surface functionalization of UCNPs based on a two-step strategy, which consists of hydrophilization with polyethyleneimine and attachment of colominic acid by electrostatic or covalent bond formation. Analysis revealed that regardless of the nature of the bond, colominic acid acted as a non-cytotoxic UCNP surface coating with low nonspecific blood protein adsorption. UCNP-colominic acid nanocomplexes exhibited low uptake by macrophages in vitro, which plays an active role in inflammatory reactions. We demonstrated the superiority of colominic acid compared to polyethylene glycol coating in terms of the prolonged circulation time in the bloodstream of small animals when injected intravenously. The colominic acid coating made it possible to prolong the UCNP circulation time up to 3 h. This led to the efficient UCNP accumulation in the inflammation site due to microvascular remodeling, accompanied by an enhanced uptake and retention effect. UCNP-assisted imaging of inflammation in the whole-body mode as well as local visualization of blood vessels were acquired in vivo. These collective findings validate the functional significance of UCNP decoration with colominic acid for their application in bioimaging.

Feasibility study of the optical imaging of a breast cancer lesion labeled with upconversion nanoparticle biocomplexes.

Innovative luminescent nanomaterials, termed upconversion nanoparticles (UCNPs), have demonstrated considerable promise as molecular probes for high-contrast optical imaging in cells and small animals. The feasibility study of optical diagnostics in humans is reported here based on experimental and theoretical modeling of optical imaging of an UCNP-labeled breast cancer lesion. UCNPs synthesized in-house were surface-capped with an amphiphilic polymer to achieve good colloidal stability in aqueous buffer solutions. The scFv4D5 mini-antibodies were grafted onto the UCNPs via a high-affinity molecular linker barstar:barnase (Bs:Bn) to allow their specific binding to the human epidermal growth factor receptor HER2/neu, which is overexpressed in human breast adenocarcinoma cells SK-BR-3. UCNP-Bs:Bn-scFv4D5 biocomplexes exhibited high-specific immobilization on the SK-BR-3 cells with the optical contrast as high as 10:1 benchmarked against a negative control cell line. Breast cancer optical diagnostics was experimentally modeled by means of epi-luminescence imaging of a monolayer of the UCNP-labeled SK-BR-3 cells buried under a breast tissue mimicking optical phantom. The experimental results were analyzed theoretically and projected to in vivo detection of early-stage breast cancer. The model predicts that the UCNP-assisted cancer detection is feasible up to 4 mm in tissue depth, showing considerable potential for diagnostic and image-guided surgery applications.

Quantitative imaging of single upconversion nanoparticles in biological tissue.

The unique luminescent properties of new-generation synthetic nanomaterials, upconversion nanoparticles (UCNPs), enabled high-contrast optical biomedical imaging by suppressing the crowded background of biological tissue autofluorescence and evading high tissue absorption. This raised high expectations on the UCNP utilities for intracellular and deep tissue imaging, such as whole animal imaging. At the same time, the critical nonlinear dependence of the UCNP luminescence on the excitation intensity results in dramatic signal reduction at (∼1 cm) depth in biological tissue. Here, we report on the experimental and theoretical investigation of this trade-off aiming at the identification of optimal application niches of UCNPs e.g. biological liquids and subsurface tissue layers. As an example of such applications, we report on single UCNP imaging through a layer of hemolyzed blood. To extend this result towards in vivo applications, we quantified the optical properties of single UCNPs and theoretically analyzed the prospects of single-particle detectability in live scattering and absorbing bio-tissue using a human skin model. The model predicts that a single 70-nm UCNP would be detectable at skin depths up to 400 µm, unlike a hardly detectable single fluorescent (fluorescein) dye molecule. UCNP-assisted imaging in the ballistic regime thus allows for excellent applications niches, where high sensitivity is the key requirement.

Background free imaging of upconversion nanoparticle distribution in human skin.

Widespread applications of nanotechnology materials have raised safety concerns due to their possible penetration through skin and concomitant uptake in the organism. This calls for systematic study of nanoparticle transport kinetics in skin, where high-resolution optical imaging approaches are often preferred. We report on application of emerging luminescence nanomaterial, called upconversion nanoparticles (UCNPs), to optical imaging in skin that results in complete suppression of background due to the excitation light back-scattering and biological tissue autofluorescence. Freshly excised intact and microneedle-treated human skin samples were topically coated with oil formulation of UCNPs and optically imaged. In the first case, 8- and 32-nm UCNPs stayed at the topmost layer of the intact skin, stratum corneum. In the second case, 8-nm nanoparticles were found localized at indentations made by the microneedle spreading in dermis very slowly (estimated diffusion coefficient, D(np) = 3-7 × 10(-12) cm(2) · s(-1)). The maximum possible UCNP-imaging contrast was attained by suppressing the background level to that of the electronic noise, which was estimated to be superior in comparison with the existing optical labels.