Dataset Information

Simplified method to perform CLARITY imaging.

ABSTRACT:

Background

Imaging methods are used widely to understand structure of brain and other biological objects. However, sample penetration by light microscopy is limited due to light scattering by the tissue. A number of methods have been recently developed to solve this problem. In one approach (SeeDB) simple procedure for clarifying brain samples for imaging was described. However, this method is not compatible with immunostaining approach as SeeDB-prepared tissue is not permeable to the antibodies. Another technique for clearing brain tissue (CLARITY) was optimized for immunochemistry, but this method technically much more demanding than SeeDB.

Results

Here we report optimized protocol for imaging of brain samples (CLARITY2). We have simplified and shortened the original protocol. Following hydrogel fixation, we cut brain tissue to 1-1.5 mm thick coronal slices. This additional step enabled us to accelerate and simplify clearing, staining and imaging steps when compared to the original protocol. We validated the modified protocol in imaging experiments with brains from line M Thy1-GFP mouse and in immunostaining experiments with antibodies against postsynaptic protein PSD-95 and striatal-specific protein DARPP32.

Conclusions

The original CLARITY protocol was optimized and simplified. Application of the modified CLARITY2 protocol could be useful for a broad range of scientists working in neurobiology and developmental biology.

SUBMITTER: Poguzhelskaya E

PROVIDER: S-EPMC4049387 | biostudies-literature | 2014 May

REPOSITORIES: biostudies-literature

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Publications

Simplified method to perform CLARITY imaging.

Poguzhelskaya Ekaterina E Artamonov Dmitry D Bolshakova Anastasia A Vlasova Olga O Bezprozvanny Ilya I

Molecular neurodegeneration 20140526

<h4>Background</h4>Imaging methods are used widely to understand structure of brain and other biological objects. However, sample penetration by light microscopy is limited due to light scattering by the tissue. A number of methods have been recently developed to solve this problem. In one approach (SeeDB) simple procedure for clarifying brain samples for imaging was described. However, this method is not compatible with immunostaining approach as SeeDB-prepared tissue is not permeable to the an ...[more]

PMID: 24885504

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Project description:To develop a simplified magnetic resonance imaging method (MRI) to assess total adipose tissue (AT) and adipose tissue free mass (ATFM) from three single MRI slices in people with overweight/obesity in order to implement body composition follow-up in a clinical research setting. Body composition of 310 participants (70 women and 240 men, age: 50.8 ± 10.6 years, BMI: 31.3 ± 5.6 kg.m-2) was assessed with 3 single slices (T6-T7, L4-L5 and at mid-thigh) MRI. Multiple regression analysis was used to develop equations predicting AT and ATFM from these three single slices. Then we implemented a longitudinal phase consisting in a 2-month exercise training program during which we tested the sensitivity of these equations in a subgroup of participants with overweight/obesity (n = 79) by comparing the exercise-induced variations between predicted and measured AT and ATFM. The following equations: total AT = - 12.74105 + (0.02919 × age) + (4.27634 × sex (M = 0, F = 1)) + (0.22008 × weight) + (26.92234 × AT T6-T7) + (23.70142 × AT L4-L5) + (37.94739 × AT mid-thigh) and total ATFM = - 33.10721 + (- 0.02363 × age) + (- 3.58052 × sex (M = 0, F = 1)) + (30.02252 × height) + (0.08549 × weight) + (11.36859 × ATFM T6-T7) + (27.82244 × ATFM L4-L5) + (58.62648 × ATFM mid-thigh) showed an excellent prediction (adjusted R2 = 97.2% and R2 = 92.5%; CCC = 0.986 and 0.962, respectively). There was no significant difference between predicted and measured methods regarding the AT variations (- 0.07 ± 2.02 kg, p = 0.70) and the ATFM variations (0.16 ± 2.41 kg, p = 0.49) induced by 2-months of exercise training. This simplified method allows a fully accurate assessment of the body composition of people with obesity in less than 20 min (10 min for images acquisition and analysis, respectively), useful for a follow-up.

Dataset Information

Simplified method to perform CLARITY imaging.

Background

Results

Conclusions

Publications

Simplified method to perform CLARITY imaging.

Similar Datasets

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