Lorenz Lechner

High-resolution helium ion microscopy of epididymal epithelial cells and their interaction with spermatozoa

We examined the rat and mouse epididymis using helium ion microscopy (HIM), a novel imaging technology that uses a scanning beam of He(+) ions to produce nanometer resolution images of uncoated biological samples. Various tissue fixation, sectioning and dehydration methods were evaluated for their ability to preserve tissue architecture. The cauda epididymidis was luminally perfused in vivo to remove most spermatozoa and the apical surface of the epithelial lining was exposed. Fixed epididymis samples were then subjected to critical point drying (CPD) and HIM. Apical stereocilia in principal cells and smaller apical membrane extensions in clear cells were clearly distinguishable in both rat and mouse epididymis using this technology. After perfusion with an activating solution containing CPT-cAMP, a permeant analog of cAMP, clear cells exhibited an increase in the number and size of membrane ruffles or microplicae. In contrast, principal cells did not exhibit detectable structural modifications. High-resolution HIM imaging clearly showed the ultrastructure of residual sperm cells, including the presence of concentric rings on the midpiece, and of cytoplasmic droplets in some spermatozoa. Close epithelium-sperm interactions were also detected. We found a number of sperm cells whose heads were anchored within the epididymal epithelium. In certain cases, the surface of the sperm cytoplasmic droplet was covered with vesicle-like structures whose size is consistent with that of epididymosomes. In conclusion, we describe here the first application of HIM technology to the study of the structure and morphology of the rodent epididymis. HIM technology represents a major imaging breakthrough that can be successfully applied to study the epididymis and spermatozoa, with the goal of advancing our understanding of their structure and function.

Advanced metrology and inspection solutions for a 3D world

Semiconductor devices and packages have firmly moved in to an era where scaling is driven by 3D architectures. However, most of the metrology and inspection technologies in use today were developed for 2D devices and are inadequate to deal with 3D structures. An additional complication is the need for specific structural and defect information that may be buried deep within a 3D structure. We present concepts and technologies that allow for 3D imaging as well as tomography, enabling engineers to view structural information with unprecedented clarity, detail and speed.

A Modern Correlative Workflow Environment to Master the Multi-scale Challenge

In most fields of study, it is imperative to understand the behavior of a system across several length scales in three dimensions in order to properly address the structural parameters that govern its performance. In order to characterize a system, be it the brain, a structural metal alloy, or a porous rock reservoir, multiple microscopic methods have evolved to specialize in capturing a relatively well defined window of scales, modalities or dimensions of information. Examples of this include medical-CT, confocal light microscopy, X-ray tomography, FIBSEM tomography, serial block face SEM, TEM tomography, atom probe tomography, and more. As these techniques have progressed individually, a clear challenge that has emerged has been how to intelligently navigate to and acquire 3D volumes of interest (from centimeter to nanometer), and, subsequently, to fuse multi-scale and multi-modality datasets in such a way that leaves the microscopist in control as recently published in the context of a corrosion study [1].