Three-dimensional characterization of cell clusters using synchrotron-radiation-based micro-computed tomography.
Micro-computed tomography with the highly intense, monochromatic X rays produced by the synchrotron is a superior method to nondestructively measure the local absorption in three-dimensional space. Because biological tissues and cells consist mainly of water as the surrounding medium, higher absorbing agents have to be incorporated into the structures of interest. Even without X-ray optics such as refractive lens, one can uncover the stain distribution with the spatial resolution of about 1 mum. Incorporating the stain at selected cell compartments, for example, binding to the RNA/DNA, their density distribution becomes quantified. In this communication, we demonstrate that tomograms obtained at the beamlines BW2 and W2 (HASYLAB at DESY, Hamburg, Germany) and 4S (SLS, Villigen, Switzerland) clearly show that the RNA/DNA-stained HEK 293 cell clusters have a core of high density and a peripheral part of lower density, which correlate with results of optical microscopy. The inner part of the clusters is associated with nonvital cells as the result of insufficient oxygen and nutrition supply. This necrotic part is surrounded by (6 +/- 1) layers of vital cells.
Müller
Bert
B
Computer Vision Laboratory ETH Zürich, Gloriastrasse 35, CH-8092 Zürich, Switzerland. bmueller@vision.ee.ethz.ch
Riedel
Marco
M
Thurner
Philipp J
PJ
D018929:Cell Culture Techniques / Q000379:methods; D002460:Cell Line; D016895:Culture Media, Serum-Free; D004247:DNA / Q000737:chemistry; D005976:Glutaral; D006801:Humans; D007668:Kidney / Q000166:cytology* / Q000648:ultrastructure; D008968:Molecular Conformation; D017785:Photons; D012313:RNA / Q000737:chemistry; D017356:Synchrotrons; D014057:Tomography, X-Ray Computed
D016895:Culture Media, Serum-Free; D012313:RNA; D004247:DNA; D005976:Glutaral