Whole-body imaging techniques play a vital role in exploring the interplay of physiological systems in maintaining health and driving disease.
We introduce wildDISCO, a new approach for whole-body immunolabeling, optical clearing and imaging in mice, circumventing the need for transgenic reporter animals or nanobody labeling and so overcoming existing technical limitations. We identified heptakis-β-cyclodextrin as a potent enhancer of cholesterol extraction and membrane permeabilization, enabling deep, homogeneous penetration of standard antibodies without aggregation.
We evaluated wildDISCO against other established methods in terms of their efficacy in staining an entire mouse using standard antibodies. In a direct side-by-side comparison using the synapsin 1 antibody, we observed incomplete staining in addition to many blurry imaged organs from iDISCO, uDISCO and PEGASOS stained samples .
Although wildDISCO has proved successful with a wide range of IgG antibodies, further validation with a broader selection of antibodies is necessary. Here, we have already listed more than 30 validated antibodies, and we will continue to update the validated antibodies regularly for the scientific community at the online website . Second, the high-resolution three-dimensional imaging generated by wildDISCO results in large amounts of data that can be challenging to analyze effectively.
For quantification of the enteric plexus in the duodenum between germ-free mice and wildtype mice, five 200 × 200 × 200 μmcubic volumes along the portal triads were randomly selected from the reconstructed 3D images in Imaris. The length of the PGP 9.5-positive enteric plexus in each cubic volume was traced using Imaris Filament Tracer.Colocalization analysis was carried out using BitPlane Imaris.
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