2D and 3D X-ray observations of ductal carcinoma in situ (DCIS) are herein reported. A large area involving a distorted linear structure and ductus lactiferi has been visualized by 2D XDFI (X-ray dark-field imaging) and furthermore a rod measuring 3.6 mm in diameter and 4.7 mm in height excised from the tissue was observed by 3D X-ray CT. These data were obtained using X-ray optics DEI (diffraction-enhanced imaging). A newly made algorithm was used for CT. The data from 900 projections at an interval of 0.2 degrees were used. As a result the ductus lactiferi and microcalcification in a 3D form were all clearly visible. The spatial resolution available was approximately 100 μm. This modality is thus considered to make a pathological diagnosis possible based on the X-ray findings.
Photodamage occurs, to a greater or lesser extent, during fluorescence bioimaging, and various procedures are empirically used to reduce the extent of this damage. However, measuring photodamage is difficult. Nitric oxide (NO), a gaseous messenger, is a free radical, and can be induced by light-exposure. In the present study, NO production was determined by the treatment of cultured cardiomyocytes, hippocampal neurons, dorsal root ganglion cells and serous peritoneal mast cells with 4, 5-diaminofluorescein (DAF-2). During laser irradiation in confocal microscopy, the fluorescence intensity of DAF-2 was increased in many punctuate regions of the cytoplasm as well as the nuclei of the cells examined. After the increase in NO levels, the beating of cardiomyocytes and the exocytosis of mast cells ceased. A stronger laser power caused a more intense peak and a more rapid increase in NO production. NO production in mast cells under various conditions was observed, in an attempt to reduce the extent of photodamage. Intermittent irradiation had no effect on NO production. Anti-oxidants (bovine serum albumin, vitamin E and melatonin), inhibited the peak and delayed the time course for NO production at a weak laser power, but had no effect on the bleaching of Indo-1, a fluorescent calcium indicator. In all bioimaging experiments in which a fluorescence technique is involved, the possibility of photodamage should be considered, and monitoring NO production by DAF-2 continues to be a relatively straightforward method for detecting photodamage.
Using the triple staining method (FDA, PI, hematoxylin) and the lumogallion staining method, the aluminum (Al) distribution and condition of the plasma membrane in cultured tobacco cells were studied. After the cells were treated with 100 mM Al at pH 4.0 for 24 h, the triple staining method was performed to simultaneously observe the Al accumulation and any damage to the plasma membrane of the cells. These result showed that no Al accumulation to be observed in the cells whose plasma membrane was not damaged. On the other hand, Al was found to be accumulated in the nuclei, plasma membrane and other organelles when the plasma membrane was damaged, thus suggesting that Al accumulation to the organelles occurred after the plasma membrane was damaged. To study the Al accumulation in more detail, the lumogallion staining method was carried out to detect any trace amounts of Al in cells. Under confocal laser microscopy, Al binding to the organelles was detected as early as after 15 min of Al treatment. The spatial Al distribution of the cells was constructed by superposing cross section images. The stereographs showed the distribution of Al on the plasma membrane not to be uniform, thus suggesting the existence of some specific Al-binding sites where Al toxicity might be induced.