The skin barrier is fundamental to terrestrial life and its evolution; it upholds homeostasis and protects against the environment. Skin barrier capacity is controlled by lipids that fill the extracellular space of the skin's surface layer--the stratum corneum. Here we report on the determination of the molecular organization of the skin's lipid matrix in situ, in its near-native state, using a methodological approach combining very high magnification cryo-electron microscopy (EM) of vitreous skin section defocus series, molecular modeling, and EM simulation. The lipids are organized in an arrangement not previously described in a biological system-stacked bilayers of fully extended ceramides (CERs) with cholesterol molecules associated with the CER sphingoid moiety. This arrangement rationalizes the skin's low permeability toward water and toward hydrophilic and lipophilic substances, as well as the skin barrier's robustness toward hydration and dehydration, environmental temperature and pressure changes, stretching, compression, bending, and shearing.
Amorphous silicon carbide nanosprings, as well as biphase (crystalline core/amorphous sheath) helical nanowires, have been synthesized by
plasma enhanced chemical vapor deposition. Both variants grow via the vapor−liquid−solid mechanism. The formation of the amorphous
silicon carbide nanosprings is explained in terms of the contact angle anisotropy model initially proposed to explain the formation of amorphous
boron carbide nanosprings. A modified contact angle anisotropy model, where the presence of temperature gradients within the catalyst are
imposed, has been proposed to explain the formation of the biphase helical nanowires. The basis for this model is that the crystalline core
acts to pin the catalyst, thereby prohibiting nanospring formation. The model is supported by the experimental observation of a transition to
nanospring growth at the point of extinction of the crystalline core of a linear biphase nanowire at a position where the catalyst is off-center
with respect to the axis of the growth direction.
Volume microscopy at high resolution is increasingly required to better understand cellular functions in the context of three-dimensional assemblies. Focused ion beam (FIB) milling for serial block face imaging in the scanning electron microscope (SEM)is an efficient and fast method to generate such volume data for 3D analysis. Here, we apply this technique at cryo-conditions to image fully hydrated frozen specimen of mouse optic nerves and Bacillus subtilis spores obtained by high-pressure freezing (HPF). We established imaging conditions to directly visualize the ultrastructure in the block face at −150°C by using an in-lens secondary electron (SE) detector. By serial sectioning with a focused ion beam and block face imaging of the optic nerve we obtained a volume as large as X=7.72 µm, Y=5.79 µm and Z=3.81 µm with a lateral pixel size of 7.5 nm and a slice thickness of 30 nm in Z. The intrinsic contrast of membranes was sufficient to distinguish structures like Golgi cisternae, vesicles, endoplasmic reticulum and cristae within mitochondria and allowed for a threedimensional reconstruction of different types of mitochondria within an oligodendrocyte and an astrocytic process. Applying this technique to dormant Bacillus subtilis spores we obtained volumes containing numerous spores and discovered a bright signal in the core, which can not be related to any known structure so far. In summary, we describe the use of cryo FIB-SEM as a tool for direct and fast 3D cryo-imaging of large native frozen samples including tissues.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.