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A guide for optimal iodine staining and high-throughput diceCT scanning in snakes
Sean Callahan1,2, Jenna M Crowe-Riddell1,3, Ramon S Nagesan1
1Museum of Zoology University of Michigan Ann Arbor MI USA.
Ecology and Evolution
|September 15, 2021
Summary
This study introduces a high-throughput workflow for diffusible iodine-based contrast-enhanced computed tomography (diceCT) scanning in snakes. The method enables detailed soft tissue visualization for ecological and evolutionary research without specimen damage.
Area of Science:
- Comparative anatomy
- Vertebrate paleontology
- Medical imaging
Background:
- Diffusible iodine-based contrast-enhanced computed tomography (diceCT) allows non-destructive soft tissue visualization in micro-CT (µCT) scans.
- Current diceCT applications are limited in taxonomic scope, hindering broad application in comparative studies.
- Standardized protocols are needed to expand diceCT use in non-model organisms like snakes.
Purpose of the Study:
- To establish best practices for diceCT staining and high-throughput µCT scanning in snakes.
- To generate a comprehensive diceCT dataset for diverse snake species.
- To assess the feasibility and applications of diceCT for ecological and evolutionary analyses in snakes.
Main Methods:
- Developed a staining and high-throughput µCT scanning workflow for 23 snake species (23 specimens).
- Scanned entire bodies and heads, generating both iodine-stained (soft tissue) and unstained (skeletal) µCT data.
- Determined optimal staining duration based on body size and assessed scan quality via grayscale values and tissue differentiation.
Main Results:
- Optimal staining time for diceCT increases linearly with snake body size, with head radius being a key predictor.
- High-quality scans exhibit clear soft tissue contrast; under/over-staining compromises data quality and segmentation.
- All generated scans provided usable data for ecological and evolutionary research, despite minor variations in staining.
- Ethanol destaining effectively removed iodine staining but required longer durations than previously reported.
Conclusions:
- The presented workflow is the first high-throughput method for full-body skeletal and diceCT scanning in snakes.
- This technique significantly increases publicly available diceCT data for reptiles and can be adapted for other elongate vertebrates.
- diceCT is a valuable tool for non-destructive anatomical studies in ecology and evolutionary biology.
