Abstract
Acquiring information on the precise distribution of a tumor is essential to evaluate intratumoral heterogeneity. Conventional hematoxylin and eosin staining, which has been used by pathologists for more than 100 years, is the gold standard of tumor diagnosis. However, it is difficult to stain entire tumor tissues with hematoxylin and eosin and then acquire the three-dimensional distribution of cells in solid tumors due to difficulties in the staining and rinsing. In this paper, we propose a modified hematoxylin and eosin staining method, in which delipidation and ultrasound waves were applied to enhance tissue permeability and accelerate dye diffusion. This improved hematoxylin and eosin staining method is termed iHE (intact tissue hematoxylin and eosin staining). We applied the iHE method to stain intact organs of mice, which were then sectioned and imaged sequentially. The results showed that the whole tissue was stained homogeneously. Combined with micro-optical sectioning tomography (MOST), the iHE method can be used for 3D volume imaging and to evaluate the intratumoral heterogeneity of the entire tumor tissue spatially. Therefore, this method may help to accurately diagnose the invasion stage of tumors and guide clinical treatments.
Highlights
For visualization of lesions in tissues, many techniques have been developed and can be divided into two categories
If an intact tumor tissue can be stained with H&E, and volume bright-field imaging can be applied to the stained sample, volume imaging can be combined with the gold standard of tumor diagnosis
Staining of tissues as large as a mouse brain is rarely performed because it is difficult to apply traditional H&E staining to intact tissue without any alterations
Summary
The stainless-steel container was wrapped using a silicone heating pad to ensure a stable solution temperature in the plastic centrifuge tubes. If blood vessel staining was needed, perfusion was performed using carbon ink solution (20% carbon ink diluted in 0.01 M PBS solution containing 40% acrylamide and 5% bis-acrylamide); azo diisopropyl imidazoline hydrochloride served as the initiator. The mouse tissues were dehydrated with 75%, 95% and absolute ethanol each for 1.5 h at 60–70 °C. The tissues were stained with Harris’ hematoxylin solution for 6 h at a temperature of 60–70 °C and were rinsed in tap water until the water was colorless. We soaked the tissue in saturated lithium carbonate solution for 12 h and rinsed it with tap water. The stained tissues were cut into 7-μm slices, dewaxed, mounted with neutral balsam and imaged using Nikon NIS-Elements microscopy
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
