抗原变异可使得多种致病微生物易于逃避宿主免疫应答。表达在感染红细胞表面的恶性疟原虫红细胞表面蛋白1（PfPMP1）与感染红细胞、内皮细胞、树突状细胞以及胎盘的单个或多个受体作用，在黏附及免疫逃避中起关键的作用。每个单倍体基因组var基因家族编码约60种成员，通过启动转录不同的var基因变异体为抗原变异提供了分子基础。

疟原虫var基因转换速率变化导致抗原变异[英]／Paul H, Robert P, Christodoulou Z, et al//Proc Natl Acad Sci U S A

X-ray crystallography has been central to the development of many fields of science over the past century. It has now matured to a point that as long as good-quality crystals are available, their atomic structure can be routinely determined in three dimensions. However, many samples in physics, chemistry, materials science, nanoscience, geology, and biology are noncrystalline, and thus their three-dimensional structures are not accessible by traditional x-ray crystallography. Overcoming this hurdle has required the development of new coherent imaging methods to harness new coherent x-ray light sources. Here we review the revolutionary advances that are transforming x-ray sources and imaging in the 21st century.

/pdf/beyond-crystallography-diffractive-imaging-using-coherent-x-3q2ojmlt88.pdf

Beyond crystallography: Diffractive imaging using coherent x-ray light sources

Very high resolution X-ray imaging has been the subject of considerable research over the past few decades. However, the spatial resolution of these methods is limited by the manufacturing quality of the X-ray optics. More recently, lensless X-ray imaging has emerged as a powerful approach that is able to circumvent this limitation. A number of classes of lensless X-ray imaging have been developed so far, with many based on other forms of optics. Here we report the key progress in this area, describe the potential applications for biology and materials science, and discuss the prospect for imaging single molecules using X-ray free-electron lasers.

/pdf/coherent-lensless-x-ray-imaging-5w0hqtqn2i.pdf

Coherent lensless X-ray imaging

Microscopy has greatly advanced our understanding of biology. Although significant progress has recently been made in optical microscopy to break the diffraction-limit barrier, reliance of such techniques on fluorescent labeling technologies prohibits quantitative 3D imaging of the entire contents of cells. Cryoelectron microscopy can image pleomorphic structures at a resolution of 3–5 nm, but is only applicable to thin or sectioned specimens. Here, we report quantitative 3D imaging of a whole, unstained cell at a resolution of 50–60 nm by X-ray diffraction microscopy. We identified the 3D morphology and structure of cellular organelles including cell wall, vacuole, endoplasmic reticulum, mitochondria, granules, nucleus, and nucleolus inside a yeast spore cell. Furthermore, we observed a 3D structure protruding from the reconstructed yeast spore, suggesting the spore germination process. Using cryogenic technologies, a 3D resolution of 5–10 nm should be achievable by X-ray diffraction microscopy. This work hence paves a way for quantitative 3D imaging of a wide range of biological specimens at nanometer-scale resolutions that are too thick for electron microscopy.

/pdf/quantitative-3d-imaging-of-whole-unstained-cells-by-using-x-3yzgb0d0yi.pdf

Quantitative 3D imaging of whole, unstained cells by using X-ray diffraction microscopy.

New diffractive imaging techniques using coherent x-ray beams have made possible nanometer-scale resolution imaging by replacing the optics in a microscope with an iterative phase retrieval algorithm However, to date very high resolution imaging (< 40nm) was limited to large-scale synchrotron facilities Here, we present a significant advance in image resolution and capabilities for desktop soft x-ray microscopes that will enable widespread applications in nanoscience and nanotechnology Using 13nm high harmonic beams, we demonstrate a record 22nm spatial resolution for any tabletop x-ray microscope Finally, we show that unique information about the sample can be obtained by extracting 3-D information at very high numerical apertures

Ultrahigh 22 nm resolution coherent diffractive imaging using a desktop 13 nm high harmonic source

We report the first image of an intact, frozen hydrated eukaryotic cell using x-ray diffraction microscopy, or coherent x-ray diffraction imaging. By plunge freezing the specimen in liquid ethane and maintaining it below −170 °C, artifacts due to dehydration, ice crystallization, and radiation damage are greatly reduced. In this example, coherent diffraction data using 520 eV x rays were recorded and reconstructed to reveal a budding yeast cell at a resolution better than 25 nm. This demonstration represents an important step towards high resolution imaging of cells in their natural, hydrated state, without limitations imposed by x-ray optics.

http://xrm.phys.northwestern.edu/research/pdf_papers/2009/huang_prl_2009.pdf

Soft X-ray diffraction microscopy of a frozen hydrated yeast cell.

Using a signal-to-noise ratio estimation based on correlations between multiple simulated images, we compare the dose efficiency of two soft x-ray imaging systems: incoherent brightfield imaging using zone plate optics in a transmission x-ray microscope (TXM), and x-ray diffraction microscopy (XDM) where an image is reconstructed from the far-field coherent diffraction pattern. In XDM one must computationally phase weak diffraction signals; in TXM one suffers signal losses due to the finite numerical aperture and efficiency of the optics. In simulations with objects representing isolated cells such as yeast, we find that XDM has the potential for delivering equivalent resolution images using fewer photons. This can be an important advantage for studying radiation-sensitive biological and soft matter specimens.

Signal-to-noise and radiation exposure considerations in conventional and diffraction x-ray microscopy

TEMGYM Advanced: Software for electron lens aberrations and parallelised electron ray tracing.


 The high detection efficiencies of direct electron detectors facilitate the routine collection of low fluence electron micrographs and diffraction patterns. Low dose and low fluence electron microscopy experiments are the only practical way to acquire useful data from beam sensitive pharmaceutical and biological materials. Appropriate modeling of low fluence images acquired using direct electron detectors is, therefore, paramount for quantitative analysis of the experimental images. We have developed a new open-source Python package to accurately model any single layer direct electron detector for low and high fluence imaging conditions, including a means to validate against experimental data through computation of modulation transfer function and detective quantum efficiency.

InFluence: An Open-Source Python Package to Model Images Captured with Direct Electron Detectors

An interactive simulation of a transmission electron microscope (TEM) called TEMGYM Basic is developed here, which enables users to understand how to operate and control an electron beam without the need to access an instrument. TEMGYM Basic allows users to familiarize themselves with alignment procedures offline, reducing the time and money required to become a proficient TEM operator. In addition to teaching the basics of electron beam alignments, the software enables users to create bespoke microscope configurations and develop an understanding of how to operate the configurations without sitting at a microscope. TEMGYM Basic also creates static ray diagram figures for a given lens configuration. The available components include apertures, lenses, quadrupoles, deflectors and biprisms. The software design uses first-order ray transfer matrices to calculate ray paths through each electron microscope component, and the program is developed entirely in Python to facilitate compatibility with machine-learning packages for future exploration of automated control.

Andrew Stewart

Papers

Soft X-ray diffraction microscopy of a frozen hydrated yeast cell.

Signal-to-noise and radiation exposure considerations in conventional and diffraction x-ray microscopy

TEMGYM Advanced: Software for electron lens aberrations and parallelised electron ray tracing.

InFluence: An Open-Source Python Package to Model Images Captured with Direct Electron Detectors

TEMGYM Basic: transmission electron microscopy simulation software for teaching and training of microscope operation