STAINING TISSUE-DERIVED MYCOBACTERIUM-LEPRAE WITH FLUORESCEIN DIACETATE AND ETHIDIUM-BROMIDE

  • 1 January 1984
    • journal article
    • research article
    • Vol. 52  (2) , 176-182
Abstract
A fluorescent staining procedure incorporating the use of fluorescein diacetate (FDA) and ethidium bromide (EB) was previously shown to accurately measure the viability of saprophytic mycobacterial cells. Green-stained cells were shown to be viable and red-stained cells dead. Staining M. leprae cells with FDA/EB was complicated by interfering tissue components which masked the presence of stained bacteria. A petroleum ether separation technique enabled 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 indicated that patients who have undergone 3 or 24 mo. 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 mo. post-infection displayed > 90% green-stained cells. There was no correlation between the FDA/EB staining method and the morphological index.

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