Protocol for Imaging and Analysis of Mouse Tumor Models with CUBIC Tissue Clearing

Abstract

Summary Tissue-clearing technologies have developed rapidly in the past decade, especially for use in neuroscience research. We previously reported that CUBIC, which is one tissue-clearing method, is useful for applications in cancer research. CUBIC cancer analysis can be used to detect cancer metastasis with single-cell resolution at whole mouse body/organ level. This approach can also analyze the tumor characteristics with high-quality 3D images. Here, we describe a detailed CUBIC cancer protocol from tissue clearing, capturing 3D images and post-immunohistochemistry. For complete details on the use and execution of this protocol, please refer to Kubota et al. (2017) .

Visualization of the cancer cell cycle by tissue‐clearing technology using the Fucci reporter system

“…In our current tissue-clearing method with CUBIC reagents (CUBIC-L and CUBIC-R (N)), a perfusion step with PBS and PFA is needed to reduce autofluorescence signals from red blood cells. 18,27,28 Tainaka et al showed that CUBIC-L reagent not only has the ability of delipidation but also a high ability to decolorize tissues, especially by removing heme. 7 Here, we showed that primary tumors in the orthotopic model with 4T1-Fucci cells were also visualized with 3D images using the CUBIC reagent (Figure 3A).…”

An analysis modality for vascular structures combining tissue-clearing technology and topological data analysis

“…Tissue clearing was performed using the CUBIC regents 10 , 12 , 13 . Mice were sacrificed and perfused with 20–30 ml phosphate buffered saline (PBS) and 20–30 ml 4% PFA (Wako) in PBS.…”

Heterogenous expression of endoglin marks advanced renal cancer with distinct tumor microenvironment fitness

“…For the detection of metastatic tumors in mouse models, the whole brain and lung were cleared by CUBIC‐L/R as previously described. 21 , 22 The brains of the mice were co‐stained with anti‐α‐smooth muscle actin (α‐SMA) antibody, as indicated in Table S1 . Images were acquired with a light‐sheet fluorescence microscopy RAPID with a ×0.63 objective.…”