Direct Ultrastructural Imaging of Macrophages Using a Novel X‐Ray Contact Microscopy

Abstract

A compact, high-resolution, laser-plasma, x-ray contact microscopy method using a table-top Nd:glass laser system has been developed. This x-ray microscopy system was applied for the observation of macrophage ultrastructures. These images were produced using proximity imaging in which a 5-ns pulse of soft x-rays with wavelengths near and inside the water windows (23A-44A) produced by the laser-plasma were absorbed by the specimen and then registered on a photo resist. The x-ray images imprinted on the photo resist were then developed and analyzed with an atomic force microscope (AFM). Mouse thioglycollate-elicited peritoneal macrophages in suspension were examined by this new x-ray microscope. The x-ray images of the macrophages were compared with those observed by conventional transmission electron microscopy (TEM). The x-ray images showed no obvious organelles, including the nucleus and endoplasmic reticulum, as can be seen with TEM, but high- and low-contrast structures caused by mass distribution of carbon were observed. Thus, using the x-ray microscopy we visualized the first x-ray images of macrophage ultrastructures. The successful x-ray imaging of macrophage ultrastructure indicates that proximity x-ray microscopy may be of value in studying physiology linked to the dynamics of a cell.

X-ray micrography and imaging of Escherichia coli cell shape using laser plasma pulsed point x-ray sources

“…In collaboration with the Center for Research and Education in Optics and Lasers at the University of Central Florida (Orlando, FL) we performed a pilot study to test the ultramicrography technique described above to obtain highresolution images of E. coli, other bacteria, and macromolecules (Kado et al, 1996;Rajyaguru et al, 1995;1996a,b).…”

Nanometer-Resolution Imaging of Living Cells Using Soft X-ray Contact Microscopy

“…The visualization of dry or hydrated human macrophages and Leydig cells demonstrated that SXCM allows the morphologies of actual intact cells to be viewed and encourages further technological development for biomedical research [45,51,[60][61][62]. This technique also offered the possibility of monitoring the morphological changes in microorganisms (e.g., upon exposure to antibiotics), as was demonstrated for the Gram-negative bacterium, Pseudomonas aeruginosa, and Candida albicans, a yeast, among others [63,64].…”

Development of a laser plasma x-ray microscope to observe live hydrated biological specimens

“…Most of the works observing biological specimens with laser plasma x-ray have been done with contact xray microscopy, which is the indirect imaging placing biological specimens onto PMMA photo resist and observing the developed photo resist after exposure with x-ray pulse. Biological cells, bacteria, and other biological specimens have been observed [1][2][3].…”