Whole-brain / body imaging with single-cell resolution

CUBIC

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Dr. Hiroki R. Ueda et al. developed a simple, efficient, and scalable brain / body clearing method and imaging / computational analysis pipeline, CUBIC (clear, unobstructed brain/body imaging cocktails and computational analysis). CUBIC offers a fluorescent-preserving, high-performance and device-free tissue clearing method based on hydrophilic Solutions (ScaleCUBIC Solutions).
It enables reproducible whole-organ and whole-body clearing and rapid 3D imaging with LSFM. CUBIC also provides processing and analysis of 3D images for extracting biological information.
Therefore, CUBIC presents a platform of whole-organ/body imaging with single-cell resolution and image informatics, which enables a wide range of users to perform experiments targeting cellular and organ layers with multiple samples..


Data provided by Dr. Etsuo A Susaki, and Dr. Hiroki R. Ueda, RIKEN

Features

  • Easy-to-use
  • No special equipment required
  • Follow up 2D H&E
  • High transparency
  • Applicable for many organs
  • Compatible with IF, FP and other fluorescent labels

Application1

CUBIC Application

Fixation Wash Preprocessing Delipidation Wash Permeabilization Clearing Imaging
Perfusion fixation + post fixation 4% PFA/PBS(-) PBS 50% Scale CUBIC-1 Solution Scale CUBIC-1 Solution Scale CUBIC-1 Solution PBS 50% Scale CUBIC-2 Solution Scale CUBIC-1 Solution Mounting**
Processing
Temperature
4 °C RT RT 37°C 37°C RT - 37°C RT RT - 37°C RT
Processing
Time
1 day 1 day 1 day 2 days 6 - 8 days 1 day 1 day 2 days

**Typically, 7:3 mixture of Mounting Solution 1 and 2 (RI ~1.49) can be used.

Brain Heart Lung Kindney Liver
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CUBIC

Figure1.Transmittance images of mouse whole organs before and after clearing with Scale CUBIC Solutions.

Brain Heart Liver
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Figure2.Transmittance images of mouse 2mm slices before and after clearing with Scale CUBIC Solutions.

Application2

CUBIC Application for fluorescent protein expression

Fixation Wash Preprocessing Delipidation Wash
Perfusion fixation + post fixation
4% PFA/PBS(-)
[pH 7.6 - 7.8]
PBS 50% Scale CUBIC-1 Solution Scale CUBIC-1 Solution Scale CUBIC-1 Solution PBS
Processing
Temperature
4 °C RT RT 37 °C 37 °C RT - 37°C
Processing
Time
1 day 1 day 1 day 2 days 6 - 8 days* 1 day
Fluorescent labels Wash Permebilization Clearing Imaging
PI(5 μg / mL) PB(10 mM) 50 % Scale CUBIC-2
Solution
Scale CUBIC-2
Solution
Mounting:**
Processing
Temperature
RT RT RT RT - 37 °C RT
Processing
Time
2 days > 1day 1 day 2 days

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Figure3. LSFM imaging of CAG-EGFP Tg Mouse (8w Male) whole brain, whole heart and dissected liver.

Application3

CUBIC Application with pathological methods

Data by S. Nojima.et al. : Scientific Reports ,7,9269 (2017) / Adapted.

Before CUBIC

After CUBIC
-------------------------------------------------------------------------------------------
0 day 2 days 3 days 4 days 6 days 7 days
Human
lung
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HE stain
Human
Lymph
node
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HE stain

Figure 4.Tissue clearing of various human organs with CUBIC
Tissue clearing and 3D imaging of deparaffinizedsamples with CUBIC

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Figure 5.Tissue clearing and 3D imaging of pathological specimens with CUBIC.

References

  • E. A. Susaki.et al. : Cell ,157(3),726(2014).
  • K. Tainaka.et al. : Cell ,159(4),911(2014).
  • E. A. Susaki.et al. : Nature Protocols ,10,1709(2015).
  • S. Nojima.et al. : Scientific Reports ,7,9269 (2017).

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