Comparison of conventional and microwave‐assisted processing of mouse retinas for transmission electron microscopy

Abstract

SummaryConventional fixation and processing of mammalian retinal tissues for transmission electron microscopic (TEM) examination is slow and produces ultrastructural artefacts in the photoreceptor cell layer. Among these artefacts are gaps between photoreceptor outer segment disc membranes and between photoreceptor cells in the region of the retina where the cell nuclei are located. A study was undertaken to determine whether a much more rapid microwave-assisted fixation and processing protocol would have an effect on the quality of ultrastructural preservation of the retina, particularly on the photoreceptor cell artefacts. The overall ultrastructural preservation of the retina was similar for the conventional and microwave-assisted techniques. However, the magnitudes of the photoreceptor artefacts were significantly reduced when microwave irradiation was used during primary fixation and processing. It is clear that, at least for the retina, employing microwave irradiation during specimen preparation for TEM results in superior ultrastructural preservation with a substantial reduction in the time required for sample preparation.

Advanced laboratory techniques for sample processing and immunolabeling using microwave radiation

“…The first reports of these technological advancements described in vivo labeling of embryos and the effect of variable power and sample temperature on embryo viability (Sanders and Gartner, 2001;Sanders, 2002). The technology was applied to tissue processing for electron microscopy (Giberson et al, 2003;Wendt et al, 2004;Forbes, 2002, 2003), decalcification (Tinling et al, 2004), immunohistochemistry (Munoz et al, 2004) and formaldehyde fixation of fresh tissue (Galvez et al, 2006). The results of this study are consistent with previous findings and further demonstrate that microwave-assisted fixation produced less cell shrinkage, excellent immunolabeling and dramatically reduced processing times.…”

Ontogenetic Changes in the Bacterial Symbiont Community of the Tropical Demosponge Amphimedon queenslandica: Metamorphosis Is a New Beginning

“…Samples were fixed and embedded using a Pelco Biowave (Ted Pella), using a protocol adapted from Wendt et al (2004). Cells were fixed in 3% glutaraldehyde in 0.1 M sodium cacodylate buffer in filtered seawater (FSW), and washed in fresh 0.1 M sodium cacodylate buffer before centrifugation at 10,000 g for 10 min.…”

Cytoplasm views from the Devonian: Observations on cells ofRhynia gwynne-vaughanii

“…Wang (2004) proposed that lightning played a role in the very rapid fixation process of plant cytoplasm in fossil plant tissues from the Cretaceous period. The author argued that the strong electric current and the related electromagnetic fields could induce a response similar to that of tissues fixed by microwaves (Giberson & Demaree 1999, Wendt et al 2004); this method is frequently used because it allows for high quality preservation in a short time.…”

“…Because in the cells studied in the present work such a deposition occurred on a molecular scale (no granules were detected under the microscope), it seems possible that the cytoplasm acted as a template in the deposition of iron oxides, allowing for a close reproduction of cytological structures. It is worth mentioning that the uranyl acetate staining of biological samples processed for transmission electron microscopy is improved and shortened by microwave irradiation (Wendt et al 2004).…”