Iodine vapor staining for atomic number contrast in backscattered electron and X‐ray imaging

Abstract

Iodine imparts strong contrast to objects imaged with electrons and X-rays due to its high atomic number (53), and is widely used in liquid form as a microscopic stain and clinical contrast agent. We have developed a simple technique which exploits elemental iodine's sublimation-deposition state-change equilibrium to vapor stain specimens with iodine gas. Specimens are enclosed in a gas-tight container along with a small mass of solid I2. The bottle is left at ambient laboratory conditions while staining proceeds until empirically determined completion (typically days to weeks). We demonstrate the utility of iodine vapor staining by applying it to resin-embedded tissue blocks and whole locusts and imaging them with backscattered electron scanning electron microscopy (BSE SEM) or X-ray microtomography (XMT). Contrast is comparable to that achieved with liquid staining but without the consequent tissue shrinkage, stain pooling, or uneven coverage artefacts associated with immersing the specimen in iodine solutions. Unmineralized tissue histology can be read in BSE SEM images with good discrimination between tissue components. Organs within the locust head are readily distinguished in XMT images with particularly useful contrast in the chitin exoskeleton, muscle and nerves. Here, we have used iodine vapor staining for two imaging modalities in frequent use in our laboratories and on the specimen types with which we work. It is likely to be equally convenient for a wide range of specimens, and for other modalities which generate contrast from electron- and photon-sample interactions, such as transmission electron microscopy and light microscopy. Microsc. Res. Tech. 77:1044–1051, 2014. © 2014 The Authors. Microscopy Research Technique published by Wiley Periodocals, Inc.

Iodine consumption and cognitive performance: Confirmation of adequate consumption

“…Iodine was discovered incidentally in the early 19th century (Rosenfeld, ). Iodine has an atomic number of 53 and atomic mass of 126.90447 amu, and it is available in the upper crust of the Earth as a trace element (Boyde, Mccorkell, Taylor, Bomphrey, & Doube, ). The variable geographic distribution of iodine is due to the effects of flooding, glaciation, and leaching into soil over time (Leung, Braverman, & Pearce, ).…”

Regulation of the Bone Vascular Network is Sexually Dimorphic

“…Samples were treated to remove bone matrix and mineral by successive application of 2N HCl and 7% chlorine sodium hypochlorite solutions, removing calcified tissue and leaving a solid PMMA cast. To enable visualization of soft tissue, the surfaces of polished blocks and casts were stained using either ammonium triiodide or iodine vapor …”

Evaluation of laser ablation microtomy for correlative microscopy of hard tissues

“…The extensive HDMPs in this case had originally been discovered by high‐contrast XMT of the whole femoral head, then retrodiscovered in an MRI data set, and this knowledge used in preparing macroslices for PMMA embedding, and the eventual demonstration by BSE SEM (Boyde et al ., ). This is a classic case where the ‘baby would have been thrown out with the bath water’ in the decalcifying fluid – if we had gone that route.…”

“…In every instance we had employed BSE SEM of topography free PMMA block surfaces. Most blocks were re‐imaged using BSE SEM after iodine staining – wet, using a triiodide solution, or dry, by iodine vapour to reveal cells and uncalcified tissue like cartilage, tendon, ligament, osteoid and muscle (Boyde, ; Boyde et al ., ,b; Ley et al ., ). Many of the samples had also been studied by confocal fluorescence and reflection imaging (Boyde et al ., ) and several by in‐house high‐contrast x‐ray microtomography (XMT: Boyde et al ., ) or standard commercial microtomography (μCT: Boyde et al ., ).…”