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Smart Biomarker-Coated PbS Quantum Dots for Deeper Near-Infrared Fluorescence Imaging in the Second Optical Window
Published in Lingyan Shi, Robert R. Alfano, Deep Imaging in Tissue and Biomedical Materials, 2017
During the past few years, significant progress has been made in the synthesis of PbS QDs for non-invasive deep-tissue imaging in the 2nd-NIR window [39–42]. Although PbS QDs contain heavy metal ions, these QDs coated with MUA, GSH and recombinant proteins do not show significant cytotoxicity [39, 40, 42]. These PbS QDs act as highly fluorescent, biocompatible probes for deep-tissue imaging. Since the surface of PbS QDs is easily modified with biomolecules such as antibodies, PbS QDs can be used as molecular probes for in vivo cellular imaging regarding inflammatory and cancer metastasis. With the aid of the light-sheet fluorescence microscopy [93] or two-photon excitation microscopy [94], the spatial resolution of the 2nd-NIR fluorescence imaging would be remarkably improved. We have demonstrated that the 2nd-NIR imaging can be applied for non-invasive tissue imaging of a lymph system, breast tumors, and brain. This imaging technique may have potential to use in pre-clinical studies of possible molecular diagnosis after septic encephalopathy (i.e., brain inflammatory syndrome) [95].
Intravital imaging of megakaryocytes
Published in Platelets, 2020
David Stegner, Katrin G. Heinze
A different approach is light sheet fluorescence microscopy (LSFM), which is a wide-field method that allows to image large specimen with high resolution. Samples are excited by a focused [93] or scanned [94] “sheet of light” while the fluorescence light is detected by a sensitive camera perpendicular to the illumination plane. Camera-based modalities are typically fast, and here, photodamage and fluorophore bleaching are additionally kept at a minimum as only the detected plane is illuminated at a time. 3D image stacks are generated by moving the sample through the light sheet. The disadvantage here is that mammalian model organisms are opaque for visible light so that light-sheet microscopy is usually performed on optically cleared samples, which excludes intravital imaging. Nevertheless, this technique offers the desired large FOV and is compatible with labeling strategies used for intravital microscopy. Thus, it is a complementary technique that can give important information on megakaryocyte distributions over the whole (intact, but not living) bone, the respective vasculature and inner bone structure [12] (Figure 4). Moreover, such the segmented objects in the whole bone image can serve as biological templates. Together with dynamics derived from intravital imaging, it is possible to perform meaningful computational simulations of bone marrow dynamics involving various cell types of interest [22]. Such realistic simulations with “true” objects derived from imaging could even interrogate situations that are not assessable by animal experiments for ethical or technical reasons.
Live imaging of single platelets at work
Published in Platelets, 2020
Karin Sadoul, Laurence Lafanechère, Alexei Grichine
Another open question is whether platelets are able to coordinate their response to divergent stimuli by cellular communication and, if so, whether this is restricted to neighboring platelets in contact with each other or whether messages can also be transmitted by mechanical tension sensing or chemical signaling between distant platelets in a clot. The synchronized ballooning of platelets in a clot supports the idea of a coordinated response [30]. Some further insights to answer this question may come from a microscopic technique called two-point microrheology, which consists in the analyses of correlations between fluctuations of intracellular structures in neighboring cells [49]. Detailed imaging of the collective behavior of platelets and their surroundings under thrombus-like conditions may benefit from new methods based on single plane illumination microscopy (SPIM) also called light-sheet fluorescence microscopy (LSFM) and soSPIM (single-objective SPIM) on the subcellular level [50,51]. These techniques combine fluorescence preserving, single plane excitation, and the high-resolution imaging at video-rate, compatible with single-molecule detection and tracking. They can also be applied to cell ensembles like organoids and blood clots. Other interesting approaches for deep-tissue high-resolution microscopy are the adaptive optics approaches, which compensate for the specimen induced optical aberrations and can retrieve the diffraction limited resolution (for review see [52]).