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Nanotechnology has brought a variety of new possibilities into biological discovery and clinical practice. In particular, nano-scaled carriers have revolutionalized drug delivery, allowing for therapeutic agents to be selectively targeted on an organ, tissue and cell specific level, also minimizing exposure of healthy tissue to drugs. In this review we discuss and analyze three issues, which are considered to be at the core of nano-scaled drug delivery systems, namely functionalization of nanocarriers, delivery to target organs and in vivo imaging. The latest developments on highly specific conjugation strategies that are used to attach biomolecules to the surface of nanoparticles (NP) are first reviewed. Besides drug carrying capabilities, the functionalization of nanocarriers also facilitate their transport to primary target organs. We highlight the leading advantage of nanocarriers, i.e. their ability to cross the blood-brain barrier (BBB), a tightly packed layer of endothelial cells surrounding the brain that prevents high-molecular weight molecules from entering the brain. The BBB has several transport molecules such as growth factors, insulin and transferrin that can potentially increase the efficiency and kinetics of brain-targeting nanocarriers. Potential treatments for common neurological disorders, such as stroke, tumours and Alzheimer's, are therefore a much sought-after application of nanomedicine. Likewise any other drug delivery system, a number of parameters need to be registered once functionalized NPs are administered, for instance their efficiency in organ-selective targeting, bioaccumulation and excretion. Finally, direct in vivo imaging of nanomaterials is an exciting recent field that can provide real-time tracking of those nanocarriers. We review a range of systems suitable for in vivo imaging and monitoring of drug delivery, with an emphasis on most recently introduced molecular imaging modalities based on optical and hybrid contrast, such as fluorescent protein tomography and multispectral optoacoustic tomography. Overall, great potential is foreseen for nanocarriers in medical diagnostics, therapeutics and molecular targeting. A proposed roadmap for ongoing and future research directions is therefore discussed in detail with emphasis on the development of novel approaches for functionalization, targeting and imaging of nano-based drug delivery systems, a cutting-edge technology poised to change the ways medicine is administered.

Multispectral optoacoustic tomography (MSOT) has recently been developed to enable visualization of optical contrast and tissue biomarkers, with resolution and speed representative of ultrasound. In the implementation described here, MSOT enables operation in real-time mode by capturing single cross-sectional images in <1 ms from living small animals (e.g., mice) and other tissues of similar dimensions. At the core of the method is illumination of the object using multiple wavelengths in order to resolve spectrally distinct biomarkers over background tissue chromophores. The system allows horizontal placement of a mouse in the imaging chamber and three-dimensional scanning of the entire body without the need to immerse the mouse in water. Here we provide a detailed description of the MSOT scanner components, system calibration, selection of image reconstruction algorithms and animal handling. Overall, the entire protocol can be completed within 15-30 min for acquisition of a whole-body multispectral data set from a living mouse.

We present a fast model-based inversion algorithm for quantitative 2-D and 3-D optoacoustic tomography. The algorithm is based on an accurate and efficient forward model, which eliminates the need for regularization in the inversion process while providing modeling flexibility essential for quantitative image formation. The resulting image-reconstruction method eliminates stability problems encountered in previously published model-based techniques and, thus, enables performing image reconstruction in real time. Our model-based framework offers a generalization of the forward solution to more comprehensive optoacoustic propagation models, such as including detector frequency response, without changing the inversion procedure. The reconstruction speed and other algorithmic performances are demonstrated using numerical simulation studies and experimentally on tissue-mimicking optically heterogeneous phantoms and small animals. In the experimental examples, the model-based reconstructions manifested correctly the effect of light attenuation through the objects and did not suffer from the artifacts which usually afflict the commonly used filtered backprojection algorithms, such as negative absorption values.

Optoacoustic tomography enables volumetric imaging with optical contrast in biological tissue at depths beyond the optical mean free path by the use of optical excitation and acoustic detection. The hybrid nature of optoacoustic tomography gives rise to two distinct inverse problems: The optical inverse problem, related to the propagation of the excitation light in tissue, and the acoustic inverse problem, which deals with the propagation and detection of the generated acoustic waves. Since the two inverse problems have different physical underpinnings and are governed by different types of equations, they are often treated independently as unrelated problems. From an imaging standpoint, the acoustic inverse problem relates to forming an image from the measured acoustic data, whereas the optical inverse problem relates to quantifying the formed image. This review focuses on the acoustic aspects of optoacoustic tomography, specifically acoustic reconstruction algorithms and imaging-system practicalities. As these two aspects are intimately linked, and no silver bullet exists in the path towards high-performance imaging, we adopt a holistic approach in our review and discuss the many links between the two aspects. Four classes of reconstruction algorithms are reviewed: time-domain (so called back-projection) formulae, frequency-domain formulae, time-reversal algorithms, and model-based algorithms. These algorithms are discussed in the context of the various acoustic detectors and detection surfaces which are commonly used in experimental studies. We further discuss the effects of non-ideal imaging scenarios on the quality of reconstruction and review methods that can mitigate these effects. Namely, we consider the cases of finite detector aperture, limited-view tomography, spatial under-sampling of the acoustic signals, and acoustic heterogeneities and losses.

Visualization of dynamic functional and molecular events in an unperturbed in vivo environment is essential for understanding the complex biology of living organisms and of disease state and progression. To this end, optoacoustic (photoacoustic) sensing and imaging have demonstrated the exclusive capacity to maintain excellent optical contrast and high resolution in deep-tissue observations, far beyond the penetration limits of modern microscopy. Yet, the time domain is paramount for the observation and study of complex biological interactions that may be invisible in single snapshots of living systems. This review focuses on the recent advances in optoacoustic imaging assisted by smart molecular labeling and dynamic contrast enhancement approaches that enable new types of multiscale dynamic observations not attainable with other bio-imaging modalities. A wealth of investigated new research topics and clinical applications is further discussed, including imaging of large-scale brain activity patterns, volumetric visualization of moving organs and contrast agent kinetics, molecular imaging using targeted and genetically expressed labels, as well as three-dimensional handheld diagnostics of human subjects.

We report an intensiometric, near-infrared (NIR) fluorescent, genetically encoded calcium ion (Ca 2+ ) indicator (GECI) with excitation and emission maxima at 678 nm and 704 nm, respectively. This GECI, designated NIR-GECO1, enables imaging of Ca 2+ transients in cultured mammalian cells and brain tissue with sensitivity comparable to currently available visible-wavelength GECIs. We demonstrate that NIR-GECO1 opens up new vistas for multicolor Ca 2+ imaging in combination with other optogenetic indicators and actuators.