X-ray micrography and imaging of Escherichia coli cell shape using laser plasma pulsed point x-ray sources.
High-resolution x-ray microscopy is a relatively new technique and is performed mostly at a few large synchrotron x-ray sources that use exposure times of seconds. We utilized a bench-top source of single-shot laser (ns) plasma to generate x-rays similar to synchrotron facilities. A 5 microlitres suspension of Escherichia coli ATCC 25922 in 0.9% phosphate buffered saline was placed on polymethylmethyacrylate coated photoresist, covered with a thin (100 nm) SiN window and positioned in a vacuum chamber close to the x-ray source. The emission spectrum was tuned for optimal absorption by carbon-rich material. Atomic force microscope scans provided a surface and topographical image of differential x-ray absorption corresponding to specimen properties. By using this technique we observed a distinct layer around whole cells, possibly representing the Gram-negative envelope, darker stained areas inside the cell corresponding to chromosomal DNA as seen by thin section electron microscopy, and dent(s) midway through one cell, and 1/3- and 2/3-lengths in another cell, possibly representing one or more division septa. This quick and high resolution with depth-of-field microscopy technique is unmatched to image live hydrated ultrastructure, and has much potential for application in the study of fragile biological specimens.
Rajyaguru
J M
JM
Department of Pediatric Infectious Diseases Research, Arnold Palmer Hospital for Children and Women, Orlando, Florida 32806, USA. 75301.3426@compuserve.com
Kado
M
M
Richardson
M C
MC
Muszynski
M J
MJ
D004926:Escherichia coli / Q000648:ultrastructure*; D007834:Lasers; D008853:Microscopy / Q000295:instrumentation / Q000379:methods*; D018625:Microscopy, Atomic Force / Q000295:instrumentation / Q000379:methods*; D010865:Pilot Projects; D014965:X-Rays