Recent advances in MicroCT have encouraged investigation of the microstructure of spiders and insects, and have led to an increasing number of studies examining neuroanatomy. MicroCT is a powerful tool, but its effectiveness depends on the preparation and scan settings of the appropriate tissue, especially soft non-bacterial tissues, such as muscles, organs and nerve tissue. Since application of MicroCT to spiders is only early stages, published protocols are often difficult to implement due to large variations in sample size. This study was launched to determine how to explain this variation. Our work is based on previous methods of imaging spider brains using MicroCT. It aims to integrate current knowledge and reduce the time spent troubleshooting appropriate methodologies, thereby facilitating future research into spiders and their central nervous system (CNS). Three different preparation and imaging techniques were based on published protocols using minor modifications using 216 spiders with Prosoma lengths ranging from 1.25 mm (small spiders) to 13.33 mm (large spiders). I’ve tested it. We compared the effectiveness of various specimen formulations, staining methods, and scan settings by classifying the quality of dorsal and lateral fine scans. It was observed that only phosphate sugar (PTA) staining agents result in complete staining of plant products and CNS, allowing CNS structures to be distinguished against small, medium and large spiders. Image averaging, increased number of projections, timing of exposure to images, and use of detector binning did not significantly affect image quality of small and large spiders, but was noise reduction. These settings, combined with a higher resolution and an aluminum filter, helped to improve the image quality of medium spiders. The suitability of methods regarding spider size, effort, chemical risk, and image quality was discussed.
Penna-Gonçalves, V., Willmott, N.J., J. Kelly, MB., Black, Jr., Lowe, EC., & Herberstein, ME. (2025). Compare microc staining and scanning methods for brain studies of spiders of different sizes. Journal of Comparative Neurology, 533(1), E70017. https://doi.org/10.1002/cne.70017
