A guide into the world of high-resolution 3D imaging: the case of soft X-ray tomography for the life sciences

Abstract

In the world of bioimaging, every choice made determines the quality and content of the data collected. The choice of imaging techniques for a study could showcase or dampen expected outcomes. Synchrotron radiation is indispensable for biomedical research, driven by the need to see into biological materials and capture intricate biochemical and biophysical details at controlled environments. The same need drives correlative approaches that enable the capture of heterologous but complementary information when studying any one single target subject. Recently, the applicability of one such synchrotron technique in bioimaging, soft X-ray tomography (SXT), facilitates exploratory and basic research and is actively progressing towards filling medical and industrial needs for the rapid screening of biomaterials, reagents and processes of immediate medical significance. Soft X-ray tomography at cryogenic temperatures (cryoSXT) fills the imaging resolution gap between fluorescence microscopy (in the hundreds of nanometers but relatively accessible) and electron microscopy (few nanometers but requires extensive effort and can be difficult to access). CryoSXT currently is accessible, fully documented, can deliver 3D imaging to 25 nm resolution in a high throughput fashion, does not require laborious sample preparation procedures and can be correlated with other imaging techniques. Here, we present the current state of SXT and outline its place within the bioimaging world alongside a guided matrix that aids decision making with regards to the applicability of any given imaging technique to a particular project. Case studies where cryoSXT has facilitated a better understanding of biological processes are highlighted and future directions are discussed.

A combination of soft X-ray and laser light sources offer 3D high content information on the native state of the cellular environment

“…Hence, treatment-induced artefacts are kept to a minimum. [1,2,6,7]. This illustrates a case study aided by correlative light and X-ray tomography that shows cryoSXT data augmented by cryoSIM data at beamline B24.…”

“…At synchrotron settings, cryoSXT data collection is fast depending on the exposure time, maximum rotation angles and angular step increase. Given the high-throughput nature of cryoSXT, it is ideal for the fast screening of biomaterials and detailed phenotyping of cell populations and their response to a variety of cues or environments [1,2,10].…”

“…Current transmission X-ray microscope (TXM) operation is performed using an X-ray energy of 500 eV which lies in the "water window" (above the carbon but below the oxygen absorption edges), whereupon carbon-rich biological structures absorb X-rays more than the oxygen-rich aqueous medium that surrounds them. This results in absorption contrast on direct detection projections reflecting the carbon-rich cellular ultrastructure [1][2][3][4][5][6].…”

“…Figure 1. Ribbon representation of the imaging pipeline at beamline B24 (middle strip) with representative data and beamline equipment associated with data collection on the top panel Figures in the top and middle strip were reproduced with permission from[1,2,6,7]. This illustrates a case study aided by correlative light and X-ray tomography that shows cryoSXT data augmented by cryoSIM data at beamline B24.…”

On-Line Fluorescence Microscopy for Identification and Imaging of Apoptotic Cell with Synchrotron-Based Soft X-ray Tomography

“…With on-line FM, researchers can easily find the sample buried in an ice layer and take the useful data without tedious post-processing. Most stations of synchrotron-based SXT around the world, such as B24@Diamond [ 18 ] and MISTRAL@ALBA [ 19 ], utilize an LED light source and filter module to build an on-line FM based on a built-in visible light microscope (VLM) system. This has been employed to find suitable samples with fluorescent markers during soft X-ray imaging.…”