Astrocytes are the most abundant cell type in the brain and are a crucial part of solving its mysteries. Originally assumed to be passive supporting cells, astrocyte's functions are now recognized to include active modulation and information processing at the neural synapse. The full extent of the astrocyte contribution to neural processing remains unknown. This is, in part, due to the lack of methods available for astrocyte identification and analysis. Existing strategies employ genetic tools like the astrocyte specific promoters glial fibrillary acidic protein (GFAP) or Aldh1L1 to create transgenic animals with fluorescently labeled astrocytes. Recently, small molecule targeting moieties have enabled the delivery of bright fluorescent dyes to astrocytes. Here, we review methods for targeting astrocytes, with a focus on a recently-developed methylpyridinium targeting moiety's development, chemical synthesis, and elaboration to provide new features like light-based spatiotemporal control of cell labeling.
We report the nuclear and optical in vitro and in vivo imaging of SKOV-3 cells by targeting HER2 with a bimodal trastuzumab conjugate. Previously, we have shown that desferrichrome derivatives provide a robust and versatile radiolabeling platform for the radioisotope zirconium-89. Here, we appended silicon-rhodamine functionalized linear desferrichrome to trastuzumab. This construct was radiolabeled and used to image cellular binding and antibody uptake in vitro and in vivo. The robust extinction coefficient of the SiR deep-red emissive fluorophore enables direct quantification of the number of appended chelators and fluorophore molecules per antibody. Subsequent radiolabeling of the multifunctional immunoconjugate with 89Zr was achieved with a 64 ± 9% radiochemical yield, while the reference immunoconjugate desferrioxamine (DFO)-trastuzumab exhibited a yield of 84 ± 9%. In vivo PET imaging (24, 48, 72, and 96 h post injection) and biodistribution experiments (96 h post injection) in HER2+ tumor bearing mice revealed no statistically significant difference of the two 89Zr-labeled conjugates at each time point evaluated. The bimodal conjugate permitted successful in vivo fluorescence imaging (96 h post injection) and subsequent fluorescence-guided, surgical resection of the tumor mass. This report details the first successful application of a fluorophore-functionalized desferrichrome derivative for targeted imaging, motivating further development and application of this scaffold as a multimodal imaging platform.
Astrocytes are the most abundant cells in the brain. They support neurons, adjust synaptic strength, and modulate neuronal signaling, yet the full extent of their functions is obscured by the dearth of methods for their visualization and analysis. Here, we report a chemical reporter that targets small molecules specifically to astrocytes both in vitro and in vivo. Fluorescent versions of this tag are imported through an organic cation transporter to label glia across species. The structural modularity of this approach will enable wide-ranging applications for understanding astrocyte biology.
Lipidated cyclopropenes serve as useful bioorthogonal reagents for imaging cell membranes due to the cyclopropene’s small size and ability to ligate with pro-fluorescent tetrazines. Previously, the lipidation of cyclopropenes required modification at the C3 position because methods to append lipids at C1/C2 were not available. Herein, we describe C1/C2 lipidation with the biologically active lipid ceramide and a common phospholipid using a cyclopropene scaffold whose reactivity with 1,2,4,5-tetrazines has been caged.
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