Chemical Probes for Visualizing Intact Animal and Human Brain Tissue

Abstract

Newly developed tissue clearing techniques can be used to render intact tissues transparent. When combined with fluorescent labeling technologies and optical sectioning microscopy, this allows visualization of fine structure in three dimensions. Gene-transfection techniques have proved very useful in visualizing cellular structures in animal models, but they are not applicable to human brain tissue. Here, we discuss the characteristics of an ideal chemical fluorescent probe for use in brain and other cleared tissues, and offer a comprehensive overview of currently available chemical probes. We describe their working principles and compare their performance with the goal of simplifying probe selection for neuropathologists and stimulating probe development by chemists. We propose several approaches for the development of innovative chemical labeling methods which, when combined with tissue clearing, have the potential to revolutionize how we study the structure and function of the human brain.

Increased Tau Phosphorylation in Motor Neurons From Clinically Pure Sporadic Amyotrophic Lateral Sclerosis Patients

“…Our findings therefore indicate that the role of phosphorylation at specific residues of tau should be the subject of future research in ALS. Additional methods including the use of recently developed fluorescent dyes and probes may also be considered to further assess ALS tissue for evidence of tau aggregation [48,49]. The positive correlation observed between cytoplasmic tau pS396 in the ALS motor cortex and TDP-43 stage is also of interest in light of recent work in a mouse model of chronic traumatic encephalopathy which found a reduction in TDP-43 pathology after elimination of phosphorylated tau [50], and the subsequent suggestion that that there may be a synergistic toxic relationship between the two proteins [51].…”

Optimization of GFP Fluorescence Preservation by a Modified uDISCO Clearing Protocol

“…uDISCO, a subsequently developed method, preserves endogenous fluorescence for months by replacing the tetrahydrofuran with tert-butanol during the dehydration step and adding diphenyl ether in BABB clearing solution (Pan et al, 2016 ). Though the fluorescence preservation achieved by uDISCO is relatively higher than that of 3DISCO, it is also associated with increased background fluorescence (Lai et al, 2017 ). When encountering samples with low-level fluorescence protein expression, the ability of uDISCO to preserve fluorescence is not good enough.…”

“…However, some solvent-based clearing methods, such as 3DISCO, quickly quench GFP (Ertürk and Bradke, 2013 ). The more recently developed uDISCO method can achieve a relatively high level of GFP fluorescence preservation over the accompanying inherently strong background fluorescence, which could decrease the signal-to-noise ratio of fluorescence images (Pan et al, 2016 ; Lai et al, 2017 ). Enhancing fluorescence preservation is an alternative way to improve image quality.…”

Ultrafast optical clearing method for three-dimensional imaging with cellular resolution

“…3D). We also examined whether FOCM was compatible with general staining methods such as immunofluorescence and chemical dye (35) without cell morphology disorganization. Here, c-fos antibody was applied to label active neurons in hippocampus ( Fig.…”