APPENDIX 455 



growing In test-tube cultures), and in the early stages of such growths 

 a low power of the microscope or a magnifying glass facilitates 

 observation. 



3. The Making of Cover-glass Preparations : (a) unstained — "the hang- 

 ing drop " ; (6) stained — single stains, e.g., gentian-violet, methylene- 

 blue, fuchsin, carbol-fuchsin, etc. ; or double stains by Gram's method, 

 by Ziehl-Neelsen's method, etc. This third part of the investigation is 

 obviously to prepare specimens for examination under the microscope.* 

 " The hanging drop " is a simple plan for securing the organisms for 

 microscopic examination in a more 

 or less natural condition. A hollow K~ 

 ground slide {i.e. a slide with a \| 

 shallow depression in it) is taken, 

 and a small ring of vaseline placed 



round the edge of the depression. v//Mi/)i/i)) iiih>,ii^n)) ,i/^ wmi i nun 

 Upon the under-side of a clean fi«. 40.-The Hanging Drop. 



cover-glass is placed a drop of dis- 

 tilled water, and this is inoculated with the smallest possible particle 

 taken from one of the colonies of the gelatine plate on the end of a 

 sterilised platinum wire. The cover-glass is then placed upon the ring 

 of vaseline, and the drop hangs into the space of the depression. Thus 

 is obtained a view of the organisms in a freely moving condition, if 

 they happen to be motile bacteria. In ordinary practice the hollow 

 slide may be dispensed with, and an ordinary slide used. 



With regard to staining, it will be undesirable here to dwell at length 

 upon the large number of methods which have been adopted. The 



* A good microscope is essential. It should have objectives of 1 inch, \, and 

 tV (oil Immersion). A white light, and proper adjustment of the substage condenser 

 and draw-tube are also necessary. A lens of tVth inch focal depth is the usual 

 power required for the study of bacteria, although in some cases a lens of a focal 

 length of -jigth inch, or even stronger, is desirable. Streptothrix actinomyces, which 

 belongs to the higher bacteria. Is better seen with a power of ^th inch than with one of 

 T^th inch. The principle of the immersion lens is the filling up of the space between 

 the lens and the cover-glass with a material whose refractive index is the same as 

 that of the lens, so that there will be no loss of illumination by the rays of light 

 passing through media of diiferent powers of refraction, while proceeding from the 

 object to the lens. The power of a microscope varies not only according to that of 

 the lens, but also according to the power of the eye-piece. Thus the magnifying 

 power of a 1-inch objective in Swift's microscope varies, according to the strength 

 of the eye-piece and to the fact that the draw-tube is closed or extended, from 'Ih 

 to 140 diameters ; a ith inch objective, from 175 to 690 diameters ; and a fi^th inch 

 objective, from 385 to 1627 diameters. As a high-power lens gives a picture which 

 has comparatively very little "depth" of focus, it is necessary to place the object 

 under examination in as nearly the same plane as possible. Hence the material 

 to be investigated should be reduced to an extremely thin film. 



The object should also be in the optical axis of the instrument, and secured in 

 position by means of the spring clips. In using the oil immersion lens, the body 

 tube of the microscope must be screwed down until the lens is in contact with the 

 oil and nearly touching the coverslip. The substage condenser must be screwed up 

 flush with the stage. The best light must be obtained by adjustment of the mirror, 

 and fine focus must be used. A skilful use of the microscope depends, of course, 

 upon an understanding of its parts and upon practice. 



