- Post no. 2 on lightsheet microscopy in neuroscience. Imaging whole cleared brains This is probably the most obvious use of lightsheet but I want to highlight something specific, read on. First, why is lightsheet uniquely suited for imaging whole cleared brains?
- - speed of acquisition. This matters when we talk about imaging centimeters3 of tissue. - low photobleaching of fluorescent labels during acquisition (only one plane is illuminated) - tissue stays intact. Thanks to long working distance objectives, you can image through a big piece of tissue.
- - creative mounting strategies tailored for whole cleared tissues. Important for fragile, expanded samples. - isotropic resolution in xyz can be achieved (multiview acquisition) - data handling is established (tiled acquisition setup, stitching, registration, fusion of stacks, visualization)
- The specific thing I want to highlight is that in the cleared brains one can detect not only proteins (stained with antibodies or tagged by a fluorescent protein) but also RNAs through in situ hybridization.
- The whole brain in situ hybridization protocol to detect RNAs expressed in different regions of the brain was optimized in the laboratory of Per Uhlen lab at Karolinska Institute in Stockholm. www.science.org/doi/10.1126/...
- One very useful application is to detect transcripts of immediate early genes. There is a wide range of stimuli that triggers expression of IEGs in neurons. Their expression is a good proxy to record which neurons were affected/activated by a stimulus, irrespective of the nature of the stimulus.Aug 8, 2025 07:25
- The movie: Dumas et al., 2024, Research Square (preprint) www.researchsquare.com/article/rs-4... Whole mouse brain in situ hybridization to detect Pitx2 mRNA (a homeodomain transcription factor, magenta) autofluorescence (gray), DISCO-based clearing, 4x detection
- Imaging done on Bruker Luxendo LCS SPIM. www.bruker.com/en/products-... There is a range of lightsheets for cleared tissue imaging. One special feature of LCS is that it is amenable to high throughput and some of our users do exactly that. You need to image 100s of mouse brains? No problem!
