Influence of external factors on the preservation of human nervous tissue for histological studies: review article

Santos, Bruno Lopes dos; Del‐Bel, Elaine; Pittella, José Eymard Homem; Tumas, Vítor

doi:10.5935/1676-2444.20140054

Cited by 4 publications

(4 citation statements)

References 19 publications

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“…The qualitative analysis of the 24-h cryoprotection protocol showed cellular osmotic shock in several regions, and the frozen block had compromised physical integrity, which was worse for the blocks cryoprotected with glycerol and DMSO. Long-time storage of larger brain specimens in sucrose cryoprotectant is usually not employed due to the likelihood of microbial contaminations (Santos et al, 2014). This is generally solved with sodium azide, often used as a preservative in the buffer solution, which aids cellular protection and microbial contamination (Minassian and Huang, 1979).…”

Section: Standardization Of Cryoprotectionmentioning

confidence: 99%

A technology platform for standardized cryoprotection and freezing of large-volume brain tissues for high-resolution histology

Kumarasami,

Verma,

Pandurangan

et al. 2023

Front. Neuroanat.

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Understanding and mapping the human connectome is a long-standing endeavor of neuroscience, yet the significant challenges associated with the large size of the human brain during cryosectioning remain unsolved. While smaller brains, such as rodents and marmosets, have been the focus of previous connectomics projects, the processing of the larger human brain requires significant technological advancements. This study addresses the problem of freezing large brains in aligned neuroanatomical coordinates with minimal tissue damage, facilitating large-scale distortion-free cryosectioning. We report the most effective and stable freezing technique utilizing an appropriate choice of cryoprotection and leveraging engineering tools such as brain master patterns, custom-designed molds, and a continuous temperature monitoring system. This standardized approach to freezing enables high-quality, distortion-free histology, allowing researchers worldwide to explore the complexities of the human brain at a cellular level. Our approach combines neuroscience and engineering technologies to address this long-standing challenge with limited resources, enhancing accessibility of large-scale scientific endeavors beyond developed countries, promoting diverse approaches, and fostering collaborations.

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Section: Standardization Of Cryoprotectionmentioning

confidence: 99%

A technology platform for standardized cryoprotection and freezing of large-volume brain tissues for high-resolution histology

Kumarasami,

Verma,

Pandurangan

et al. 2023

Front. Neuroanat.

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“…The current study has limitations. TSE image contrast and ex vivo brain relaxation parameters may be altered compared to the in vivo brain by agonal pathophysiology (Dos Santos et al, 2014); postmortem interval (Shepherd et al, 2009a); procurement-associated brain distortion, cuts or intraventricular air; formaldehyde fixation (Shepherd et al, 2009b); and imaging at room temperature (Ruder et al, 2012). We empirically observed that relaxation time above 2 s only increased scan duration (an undesirable result), but did not visibly affect contrast via T1 relaxation mechanisms.…”

Section: Discussionmentioning

confidence: 64%

Inner SPACE: 400-Micron Isotropic Resolution MRI of the Human Brain

et al. 2020

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Results: This MRI microscopy protocol provided excellent contrast resolution of small nuclei and internal myelinated pathways within the basal ganglia, thalamus, brainstem, and cerebellum. Contrast was sufficient to visualize the presence and variation of horizontal layers in the cerebral cortex. 3D isotropic resolution datasets facilitated simultaneous multi-planar visualization and efficient production of specific tailored oblique image orientations to improve understanding of complex neuroanatomy. Conclusion: We created an unlabeled high-resolution digital 3D MRI dataset of neuroanatomy as an online resource for readers to download, manipulate, annotate and use for clinical practice, research, and teaching that is complementary to traditional histology-based atlases. Digital MRI contrast is quantifiable, reproducible across brains and could help validate novel MRI strategies for in vivo structure visualization.

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“…The postmortem preservation of brain tissue is influenced by external factors, such as premortem and postmortem conditions and the conservation methods (fixation and freezing). Strict control of these variables is critical for good results in histochemistry and immunohistochemistry [ 20 ]. The processes of postmortem degradation can result in structural damages, changes in pH, cerebral edema, alterations of structural epitopes and in the chemical integrity of matrix components, and synapsis [ 5 , 21 , 22 ].…”

Section: Discussionmentioning

confidence: 99%

Effect of Postmortem Degradation on the Preservation of Viral Particles and Rabies Antigens in Mice Brains. Light and Electron Microscopic Study

Monroy-Gómez

Santamaría

Sarmiento

et al. 2020

Viruses

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Rabies diagnosis is mainly made on fresh brain tissue postmortem by means of the direct immunofluorescence test. However, in some cases, it is not possible to use this technique, given that the affected nervous tissue goes through a postmortem degradation process, due to problems in the handling and transport of the samples. For this reason, the preservation in time of the rabies virus inclusions was assessed, as well as the immunoreactivity and the ultrastructure of viral particles in tissue with postmortem degradation. Brains of mice inoculated with rabies virus and control mice were processed for conventional histology, immunohistochemistry, electron microscopy, and immunoelectron microscopy in different postmortem times. In the processed tissues for hematoxylin and eosin, the presence of eosinophilic inclusions was not observed beyond 12 h postmortem. Surprisingly, the immunoreactivity of the viral antigens increased with time, at least until 30 h postmortem. It was possible to easily recognize the viral particles by means of conventional electron microscopy until 12 h postmortem. Immunoelectron microscopy allowed us to identify the presence of viral antigens disseminated in the neuronal cytoplasm until 30 h postmortem, but immunoreactive viral particles were not observed. The rabies infection did not cause histological or ultrastructural alterations different from those in the control group as a result of the postmortem degradation. In conclusion, the immunohistochemistry is a reliable test for rabies diagnosis in samples with postmortem degradation and that have been fixed with aldehydes.

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Influence of external factors on the preservation of human nervous tissue for histological studies: review article

Cited by 4 publications

References 19 publications

A technology platform for standardized cryoprotection and freezing of large-volume brain tissues for high-resolution histology

A technology platform for standardized cryoprotection and freezing of large-volume brain tissues for high-resolution histology

Inner SPACE: 400-Micron Isotropic Resolution MRI of the Human Brain

Effect of Postmortem Degradation on the Preservation of Viral Particles and Rabies Antigens in Mice Brains. Light and Electron Microscopic Study

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