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The infiltration of immune cells into sites of inflammation is one key feature of immune mediated inflammatory diseases. A detailed assessment of the in vivo dynamics of relevant cell subtypes could booster the understanding of this disease and the development of novel therapies. We show in detail how advanced X-ray fluorescence imaging enables such quantitative in vivo cell tracking, offering solutions that could pave the way beyond what other imaging modalities provide today. The key for this achievement is a detailed study of the spectral background contribution from multiple Compton scattering in a mouse-scaled object when this is scanned with a monochromatic pencil X-ray beam from a synchrotron. Under optimal conditions, the detection sensitivity is sufficient for detecting local accumulations of the labelled immune cells, hence providing experimental demonstration of in vivo immune cell tracking in mice. [ABSTRACT FROM AUTHOR]

Bioluminescence imaging (BLI) is a widely used technique in preclinical scientific research, particularly in the development of mRNA-based medications and non-invasive tumor diagnostics. It has become an essential tool in current science. However, the current state of bioluminescence imaging is primarily qualitative, making it challenging to obtain quantitative measurements and to draw accurate conclusions. This fact is caused by the unique properties of optical photons and tissue interactions. In this paper, we propose an experimental setup and Geant4-simulations to gain a better understanding of the optical properties and processes involved in bioluminescence imaging. Our goal is to advance the field towards more quantitative measurements. We will discuss the details of our experimental setup, the data we collected, the outcomes of the Geant4-simulations, and additional information on the underlying physical processes. [ABSTRACT FROM AUTHOR]

X-ray fluorescence imaging (XFI) is a non-invasive detection method of small quantities of elements, which can be excited to emit fluorescence x-ray photons upon irradiation with an incident x-ray beam. In particular, it can be used to measure nanoparticle uptake in cells and tissue, thus making it a versatile medical imaging modality. However, due to substantially increased multiple Compton scattering background in the measured x-ray spectra, its sensitivity severely decreases for thicker objects, so far limiting its applicability for tracking very small quantities under in-vivo conditions. Reducing the detection limit would enable the ability to track labeled cells, promising new insights into immune response and pharmacokinetics. We present a synchrotron-based approach for reducing the minimal detectable marker concentration by demonstrating the feasibility of XFI for measuring the yet inaccessible distribution of the endogenous iodine in murine thyroids under in-vivo conform conditions. This result can be used as a reference case for the design of future preclinical XFI applications as mentioned above. [ABSTRACT FROM AUTHOR]

Simple Summary: One novel approach in cancer therapy is the use of genetically modified immune cells that are more specifically directed to a tumor than common chemotherapy. This creates the need for medical imaging methods that can be used to track these immune cells during therapy. Our study provides computer simulations of potential applications of X-ray fluorescence imaging for this purpose. We showed that if immune cells were labeled with gold nanoparticles as an imaging marker, the amounts of immune cells that would be expected to be found in a tumor or inflammation site could be detected with our setup. Our feasibility study thus shows results that are promising estimates on what can be achieved. The growing field of cellular therapies in regenerative medicine and oncology calls for more refined diagnostic tools that are able to investigate and monitor the function and success of said therapies. X-ray Fluorescence Imaging (XFI) can be applied for molecular imaging with nanoparticles, such as gold nanoparticles (GNPs), which can be used in immune cell tracking. We present a Monte Carlo simulation study on the sensitivity of detection and associated radiation dose estimations in an idealized setup of XFI in human-sized objects. Our findings demonstrate the practicability of XFI in human-sized objects, as immune cell tracking with a minimum detection limit of 4.4 × 105 cells or 0.86 μg gold in a cubic volume of 1.78 mm3 can be achieved. Therefore, our results show that the current technological developments form a good basis for high sensitivity XFI. [ABSTRACT FROM AUTHOR]

Immunotherapy has been a breakthrough in cancer treatment, yet only a subgroup of patients responds to these novel drugs. Parameters such as cytotoxic T-cell infiltration into the tumor have been proposed for the early evaluation and prediction of therapeutic response, demanded for non-invasive, sensitive and longitudinal imaging. We have evaluated the feasibility of X-ray fluorescence imaging (XFI) to track immune cells and thus monitor the immune response. For that, we have performed Monte Carlo simulations using a mouse voxel model. Spherical targets, enriched with gold or palladium fluorescence agents, were positioned within the model and imaged using a monochromatic photon beam of 53 or 85 keV. Based on our simulation results, XFI may detect as few as 730 to 2400 T cells labelled with 195 pg gold each when imaging subcutaneous tumors in mice, with a spatial resolution of 1 mm. However, the detection threshold is influenced by the depth of the tumor as surrounding tissue increases scattering and absorption, especially when utilizing palladium imaging agents with low-energy characteristic fluorescence photons. Further evaluation and conduction of in vivo animal experiments will be required to validate and advance these promising results. [ABSTRACT FROM AUTHOR]

Trace elements, functionalized nanoparticles and labeled entities can be localized with sub-mm spatial resolution by X-ray fluorescence imaging (XFI). Here, small animals are raster scanned with a pencil-like synchrotron beam of high energy and low divergence and the X-ray fluorescence is recorded with an energy-dispersive detector. The ability to first perform coarse scans to identify regions of interest, followed by a close-up with a sub-mm X-ray beam is desirable, because overall measurement time and X-ray dose absorbed by the (biological) specimen can thus be minimized. However, the size of X-ray beams at synchrotron beamlines is usually strongly dependent on the actual beamline setup and can only be adapted within specific pre-defined limits. Especially, large synchrotron beams are non-trivial to generate. Here, we present the concept of graphite-based, convex reflection optics for the one-dimensional enlargement of a 1 mm wide synchrotron beam by a factor of 5 to 10 within a 1 m distance. Four different optics are tested and characterized and their reflection properties compared to ray tracing simulations. The general shape and size of the measured reflection profiles agree with expectations. Enhancements with respect to homogeneity and efficiency can be expected with improved optics manufacturing. A mouse phantom is used for a proof-of-principle XFI experiment demonstrating the applicability of coarse and fine scans with the suggested optics design. [ABSTRACT FROM AUTHOR]

Quantitative cellular in vitro nanoparticle uptake measurements are possible with a large number of different techniques, however, all have their respective restrictions. Here, we demonstrate the application of synchrotron-based X-ray fluorescence imaging (XFI) on prostate tumor cells, which have internalized differently functionalized gold nanoparticles. Total nanoparticle uptake on the order of a few hundred picograms could be conveniently observed with microsamples consisting of only a few hundreds of cells. A comparison with mass spectroscopy quantification is provided, experimental results are both supported and sensitivity limits of this XFI approach extrapolated by Monte-Carlo simulations, yielding a minimum detectable nanoparticle mass of just 5 pg. This study demonstrates the high sensitivity level of XFI, allowing non-destructive uptake measurements with very small microsamples within just seconds of irradiation time. [ABSTRACT FROM AUTHOR]