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Phosphotungstic acid-enhanced microCT: Optimized protocols for embryonic and early postnatal mice
Kate M Lesciotto1, Susan M Motch Perrine1, Mizuho Kawasaki1
1Department of Anthropology, Pennsylvania State University, University Park, Pennsylvania.
Background:
Given the need for descriptive and increasingly mechanistic morphological analyses, contrast-enhanced microcomputed tomography (microCT) represents perhaps the best method for visualizing 3D biological soft tissues in situ. Although staining protocols using phosphotungstic acid (PTA) have been published with beautiful visualizations of soft tissue structures, these protocols are often aimed at highly specific research questions and are applicable to a limited set of model organisms, specimen ages, or tissue types. We provide detailed protocols for micro-level visualization of soft tissue structures in mice at several embryonic and early postnatal ages using PTA-enhanced microCT.
Results:
Our protocols produce microCT scans that enable visualization and quantitative analyses of whole organisms, individual tissues, and organ systems while preserving 3D morphology and relationships with surrounding structures, with minimal soft tissue shrinkage. Of particular note, both internal and external features of the murine heart, lungs, and liver, as well as embryonic cartilage, are captured at high resolution.
Conclusion:
These protocols have broad applicability to mouse models for a variety of diseases and conditions. Minor experimentation in the staining duration can expand this protocol to additional age groups, permitting ontogenetic studies of internal organs and soft tissue structures within their 3D in situ position.
Insights
Detailed protocols for phosphotungstic acid-enhanced microCT (microCT) enable high-resolution 3D soft tissue visualization in embryonic and early postnatal mice. This method preserves morphology for broad applications in developmental biology and disease modeling.
Area of Science:
- Developmental Biology
- Biomedical Imaging
- Morphology
Background:
- Microcomputed tomography (microCT) is crucial for 3D soft tissue analysis.
- Existing phosphotungstic acid (PTA) staining protocols are often limited in scope.
- A need exists for versatile PTA-enhanced microCT protocols for diverse biological samples.
Purpose of the Study:
- To provide detailed, adaptable protocols for PTA-enhanced microCT.
- To enable micro-level visualization of soft tissues in mice across developmental stages.
- To facilitate 3D morphological analysis of biological structures in situ.
Main Methods:
- Development and refinement of PTA staining protocols for microCT.
- Application of protocols to embryonic and early postnatal mouse specimens.
- High-resolution microCT scanning of whole organisms and specific tissues.
Main Results:
- Successful high-resolution 3D visualization of murine soft tissues, including heart, lungs, and liver.
- Preservation of intricate 3D morphology and spatial relationships with minimal shrinkage.
- Enabling of both qualitative and quantitative analyses of internal and external features.
Conclusions:
- The developed protocols offer broad applicability for mouse models in various research areas.
- Adaptability for different age groups allows for ontogenetic studies of soft tissues.
- Provides a valuable tool for mechanistic morphological analyses in developmental and disease research.
