220 CULTIVATION AND REPRODUCTION OF YEAST. 



Carlsburg laboratory. A small portion of the sample (previously 

 reinvigorated, if necessary) is placed in sterilised water or 

 preferably in a 0.5 per cent, solution of common salt and shaken 

 up so as to separate the agglomerations of yeast and distribute 

 the cells as uniformly as possible. The number of cells in a 

 single drop of the diluted mass having been examined under the 

 microscope, sufficient is transferred to a flask, already charged 

 with 20-100 c.c. of liquefied wort-gelatin (or must-gelatin) to 

 ensure that a tiny drop of this latter contains only a very few 

 cells. To prepare a miniature plate culture, a large cover-glass is 

 sterilised, by passing it through a spirit flame, and one side of 

 the same is then coated with a layer (about 0.2 mm. thick) of the 

 inoculated gelatin, by means of a small platinum loop, in such a 

 manner as to leave an outer ring of clear glass several milli- 

 metres wide. The cover-glass is then laid on the vaseline-coated 

 ring (c) of a sterilised Botteher cell (Fig. 159), the bottom of 

 which has already been coated with a minute quantity of sterilised 

 water (d). If this moist cell be laid horizontally on a suitable 

 foundation and exposed to a medium room temperature (i5C.), 

 the gelatin film (b) on the inside will quickly set evenly. A 

 hollow-ground microscope slide may also be used in place of the 

 Bottcher cell. It is advisable to make several of these plate cultures 



and not merely a single 

 one. As soon as the 

 gelatin film has set, the 

 cell is examined under 

 the microscope, prefer - 



FIG j ably one fitted with a 



Buttcher Cell in vertical section. Slightly nose-piece to enable the 



reduced. (After Hansen.~) two requisite objectives 



of different power (e.g., 



Zeiss Nos. A and D) to be rapidly exchanged. The low power 

 (about 40-60) is used for systematically examining the gelatin 

 film all over, to find suitable cells that are isolated and far 

 enough away from all others. The higher power (about 250- 

 400) is then used to make sure, and the exact position of the 

 selected cells is noted down. The various methods and appliances 

 for fixing the position of the cells in single-cell culture have been 

 critically reviewed by H. WILL (XXIV.) on the basis of his wide 

 experience. The author finds the simplest and most convenient 

 method and much less troublesome to the beginner than the 

 object-marker is that of marking the position of the cell direct 

 on the glass (the low power being used so that the objective is 

 about i cm. away from the slide) by means of two fine dots of ink 

 or preferably black varnish, applied to the top surface of the cover- 

 glass by means of a fine drawing-pen or a pointed inoculating 

 hook. About half a dozen or more cells are marked off in this 

 way, a necessary precaution, because it often happens that one or 



