The development of fluorescence microscopy for tubercle bacilli and its use as an adjunct to histological routine

The microscopic demonstration of the tubercle baciilus by the Ziehl-Neelsen technique has always entailed heavy demands on the time and concen-tration of the observer, but in recent years fluorescence microscopy has offered a means of greatly facilitating this task. The spontaneous fluorescence of M. tuberculosis under ultra-violet microscopy was first observed by Kaiserling (1917), but the use of fluorochrome staining is much more recent (Hagemann, 1937), and it is with this latter technique that this paper is concerned. The essential requirements are a high intensity light source and an appropriate system of filters. A blue filter eliminates the long wavelengths. The short wavelength rays elicit fluorescence from the stained preparation, and an orange eyepiece filter then allows only the fluorescence to reach the observer. The bacilli are seen as bright rods against a dark background, and are easily seen with low magnifications, permitting a much more rapid survey than with oil immersion objectives. For staining, Hagemann first employed berberine sul-phate, but later he reported superior results with auramine 0 (Hagemann, 1938), and this has be-come the standard fluorochrome for M. tubercu-losis. In reviewing the literature it is proposed to con-sider (a) the apparatus used, (b) the staining methods, and (c) the practical applications and results, and to comment on these before detailing the author's investigations. Apparatus The equipment used by Hagemann, incorpor-ating an arc lamp and quartz lenses, was not generally available, but in 1941 a number of authors reported that auramine gave adequate fluorescence with a low-voltage/high-amperage filament lamp or a projection lamp, and a norma