02054nas a2200277   4500000000100000008004100001260001300042653001200055653001300067653001700080653001100097653001200108653000900120653002500129653002200154653000900176653002600185100001400211700001400225700001500239245009800254300001100352490000700363520139200370022001401762       1984                            d  c1984 Jun10aAnimals10aEthidium10aFluoresceins10aHumans10aleprosy10aMice10aMycobacterium leprae10aPhenylenediamines10aSkin10aStaining and Labeling1 aKvach J T1 aMunguia G1 aStrand S H00aStaining tissue-derived Mycobacterium leprae with fluorescein diacetate and ethidium bromide.  a176-820 v523 a<p>A fluorescent staining procedure incorporating the use of fluorescein diacetate (FDA) and ethidium bromide (EB) has previously been shown to accurately measure the viability of saprophytic mycobacterial cells. Green-stained cells were shown to be viable and red-stained cells, dead. Staining Mycobacterium leprae cells with FDA/EB, however, was complicated by interfering tissue components which masked the presence of stained bacteria. A petroleum ether separation technique enables M. leprae to be segregated from armadillo liver tissue components and permitted M. leprae to be stained qualitatively equal to the saprophytic mycobacteria. An alternative and technically simpler method of staining M. leprae from human skin biopsies and mouse foot pads was developed which permitted the initiation of a clinical assessment of the staining method. Preliminary data indicate that patients who have undergone three or 24 months of chemotherapy possess a significantly lower percentage of green-stained M. leprae in their tissues than untreated patients. This would be expected if the FDA/EB staining method was providing an accurate measure of viability. M. leprae cells obtained from mouse foot pads which were harvested 5-13 months post-infection displayed more than 90% green-stained cells. There was no correlation between the FDA/EB staining method and the morphological index.</p>  a0148-916X