Seeing space: visualization and quantification of plant leaf structure using X-ray micro-computed tomography.

A mechanistic understanding of plant physiological processes occurring in the above-ground organs, such as photosynthesis in leaves, requires knowledge of the structural properties of the tissues, including leaf thickness (Hanba et al., 1999; Zhu et al., 2010; Niinemets et al., 2011), cell size, and stomatal distribution influencing gas exchange (Sage and Sage, 2009; Gong et al., 2011; Terashima et al., 2011). However, previous work has been unable to determine precisely how the three dimensional (3D) complexity of leaf structure impacts upon photosynthetic activity, partly due to a lack of appropriate methods for undisturbed visualization and quantification of inner structures at an appropriate high resolution (i.e. 5–10 μm). Although there is a wide range of traditional, optical-based techniques for visualization and quantification of plant morphology and anatomy, ranging from imaging of thin sections to confocal microscopy of intact specimens, their use is usually limited by the type and size of sample material. Furthermore, analysis is often time consuming, destructive, and constrained to two dimensions (2D) (classical histology) or limited in optical depth (confocal microscopy).