Imaging techniques for observing laminar geometry in the feather shaft cortex

Bird feather shafts are light, stiff and strong, but the fine details of how their structure, mechanics and function relate to one another remains poorly understood. The missing piece in our understanding may be the various fibrous layers that make up the shaft's cortex. Detailed imaging techniques are needed to enable us to capture, analyse and quantify these layers before we can begin to unravel the relationship between their structure, mechanics and function. We show that Serial-Block-Face scanning electron microscopy, scanning confocal polarised microscopy and synchrotron-based computed tomography are three suitable techniques to investigate layer thickness and fibre orientation in the feather cortex. These techniques and other are discussed in terms of their ability to resolve the fibrous laminar structure of the feather cortex, on sample preparation, and on throughput. Annotated images are presented for each and less suitable techniques are presented in the Supplementary Material. LAY DESCRIPTION: Bird feathers have a light, stiff and strong central shaft. However, the fine details of how their structure, mechanics and function relate to one another remains poorly understood. The missing piece in our understanding may have to do with how fibrous layers within the shaft vary in thickness and alignment. Detailed imaging techniques are needed so that we can quantify some of this variation before we can revisit some long-unanswered questions about the feather shaft's structure, mechanics and function. We investigate a number of microscopy techniques and show that three techniques are suitable for the sort of investigation that is required. These techniques and others are discussed in terms of their ability to resolve the layers' thickness and alignment, on sample preparation, and on the sample sizes they are able to process. Annotated images are presented and discussed for each of the three techniques and unsuitable techniques receive the same examination in the Supplementary Material.