• 1 EXPERIMENTAL DETAILS
  • 2 TISSUE CLEARING DETAILS
  • 3 SELECT PROTOCOL
  • 4 PROTOCOL MODIFICATIONS
  • 5 IMAGING and ANALYSIS
  • 6 General Questions
Specimen type
Relative age of specimen before harvest
Tissue type
Do you want to provide details on a tissue clearing protocol that was successful or unsuccessful?

If successful, how satisfied are you with this tissue clearing protocol?
Why was the tissue clearing protocol chosen?

Are you still searching for more effective tissue clearing protocols?

List of Abbreviations

Protocol
Step		Tissue Clearing Protocols
Citation Citation or common name of protocol mostly closely followed SCALE                 
REF REF: Hama H, Kurokawa H, Kawano H, et al. Scale: a chemical approach for fluorescence imaging and reconstruction of transparent mouse brain. Nat Neurosci. 2011;14(11):1481-1488. Published 2011 Aug 30. doi:10.1038/nn.2928
CUBIC        
REF REF: Tainaka K, Kubota SI, Suyama TQ, et al. Whole-body imaging with single-cell resolution by tissue decolorization. Cell. 2014;159(4):911-924. doi:10.1016/j.cell.2014.10.034
PACT        
REF REF: Yang, B. et al. Single-cell phenotyping within transparent intact tissue through whole-body clearing. Cell 158, 945–958, doi: 10.1016/j.cell.2014.07.017 (2014).
SWITCH    
REF REF: Murray E, Cho JH, Goodwin D, et al. Simple, Scalable Proteomic Imaging for High-Dimensional Profiling of Intact Systems. Cell. 2015;163(6):1500-1514. doi:10.1016/j.cell.2015.11.025
EXPANSION    
REF REF: Chen, F. et al. Expansion Microscopy. Science 347, 543-548, doi: 10.1126/science.1260088
iDISCO
REF REF:Renier N, Wu Z, Simon DJ, Yang J, Ariel P, Tessier-Lavigne M. iDISCO: a simple, rapid method to immunolabel large tissue samples for volume imaging. Cell. 2014;159(4):896-910. doi:10.1016/j.cell.2014.10.010
BABB            
REF REF:Dodt HU, Leischner U, Schierloh A, et al. Ultramicroscopy: three-dimensional visualization of neuronal networks in the whole mouse brain. Nat Methods. 2007;4(4):331-336. doi:10.1038/nmeth1036
Other, Specify        
Fixation Fixation - Type of fixative
PFA, 4% overnight
Image reference:
PFA, 4% overnight
Image reference:
PFA, 4% overnight
Image reference:
GA, 2%
overnight
Image reference:
PFA, 4% overnight
Image reference:
PFA,
4% overnight
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PFA, 4% overnight
Image reference:
Image reference:
Pre-treatment Pre-treatment Additional bleaching or tissue stabilization
Collagenase
Image reference:
Quadrol
for decolorizing
Image reference:
Hydrogel
embedding
Image reference:
Tissue gel,
low pH
Image reference:
Hydrogel
+ SA
Image reference:
Methanol dehydration for decolorizing
Image reference:
Hydrogen peroxide for decolorizing
Image reference:

Image reference:
Labeling Labeling - Antibody, dye, or fluorescence retention
Retention
Image reference:
Thy-1-YFP retention
Image reference:
Thy1-YFP retention
Image reference:
Antibody,
Tuj1
Image reference:
Passive
antibody
Image reference:
Nanobody
Image reference:
Antibody-
anti-GFP
Image reference:

Image reference:
Delipidation Delipidation - Details of lipid removal step
Urea, triton
Image reference:
Amino-
alcohol
Image reference:
Passive,SDS
Image reference:
Active electrophoresis, SDS
Image reference:
Enzyme mediated
Image reference:
THF,
Dibenzylether,
DCM
Image reference:
Methanol gradient
Image reference:

Image reference:
Refractive index matching Refractive index matching - Medium used for refractive index matching
Urea,water,
1.40
Image reference:
Aqueous: Amino-alcohol, 1.45
Image reference:
Aqueous: Histodenz, refractive index 1.46
Image reference:
Aqueous: 90% glycerol,refractive index 1.46
Image reference:
Water,
salts,refractive index 1.33-.140
Image reference:
Organic:
Dibenzyl ether, refractive index 1.54
Image reference:
Organic:
BABB,
refractive index 1.56
Image reference:

Image reference:
 
Image reference:Techniques based on hyperhydrating solutions 		Image reference:Techniques based on tissue transformation
Image reference:Techniques based on organic solvents 		Image reference:Other
Protocol Steps Protocol Enter Protocol modifications or additional details for “other” Enter citation or reference for any modifications or “Other” step (if available)
Fixation
Pre-treatment
Labeling
Delipidation
Refractive index matching
Type of analysis done on feature of interest
Level of analysis done on the image
Were the images acquired of sufficient quality to carry out the analysis?
If not, do you believe the microscope or clearing were the cause of the poor images?
In addition to the details gathered here, are there other things you would like to share about your tissue clearing experiments?

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