Rajyaguru Jayshree M, Kado Masataka, Nekula Kerry, Richardson Martin C, Muszynski Michael J
Department of Pediatric Infectious Diseases Research, Arnold Palmer Hospital for Children and Women,85 West Miller Street, Suite 303, Orlando, FL 32806,USA.
Center for Research and Education in Optics and Lasers, University of Central Florida,4000 Central Florida Blvd, Orlando, FL 32826-2700,USA.
Microbiology (Reading). 1997 Mar;143 ( Pt 3):733-738. doi: 10.1099/00221287-143-3-733.
Electron microscopy is still the most frequently used method for visualization of subcellular structures in spite of limitations due to the preparation required to visualize the specimen, High resolution X-ray microscopy is a relatively new technique, still under development and restricted to a few large synchrotron X-ray sources. We utilized a single-shot laser (nanosecond) plasma to generate X-rays similar to synchrotron facilities to image live cells of Candida albicans. The emission spectrum was tuned for optimal absorption by carbon-rich material. The photoresist was then scanned by an atomic force microscope to give a differential X-ray absorption pattern. Using this technique, with a sample image time of 90 min, we have visualized a distinct 152.24 nm thick consistent ring structure around cells of C albicans representing the cell wall, and distinct 'craters' inside, one of 570-90 nm diameter and three smaller ones, each 400 nm in diameter. This technique deserves further exploration concerning its application in the ultrastructural study of live, hydrated microbiological samples and of macromolecules.
尽管由于观察标本所需的制备过程存在局限性，但电子显微镜仍是观察亚细胞结构最常用的方法。高分辨率X射线显微镜是一种相对较新的技术，仍在发展中，并且仅限于少数大型同步加速器X射线源使用。我们利用单次激光（纳秒级）等离子体产生类似于同步加速器设备产生的X射线，以对白色念珠菌的活细胞进行成像。发射光谱经过调整，以实现富含碳的物质的最佳吸收。然后用原子力显微镜扫描光刻胶，以给出差分X射线吸收图案。使用该技术，在90分钟的样品成像时间内，我们观察到白色念珠菌细胞周围有一个明显的、厚度为152.24 nm的连续环形结构，代表细胞壁，内部还有明显的“坑”，其中一个直径为570 - 90 nm，另外三个较小，每个直径为400 nm。这项技术在活的、含水的微生物样品和大分子的超微结构研究中的应用值得进一步探索。
