276 Profs. Percy Frankland and Marshall Ward. 



used previously in isolating the yeasts and bacteria of the " ginger- 

 beer plant."* 



A small drop of nutrient gelatine or of broth, properly infected 

 with ripe spores from an agar tube, was placed on the under surface 

 of a sterilised cover-slip, and the latter luted by means of sterile 

 gelatine, so as to serve as the lid to one of the glass cultivating 

 chambers, properly sterilised, the floor of which consisted of a glass 

 slip, luted by means of melted paraffin. Over the floor of the cham- 

 ber was spread a layer of sterilised distilled water, and the tubular 

 arms were plugged with cotton- wool saturated with water, all pro- 

 perly sterilised. The hanging drop, selected by examining a number 

 and choosing that which appeared to contain only one, or at most a 

 very few spores, separated by a sufficient distance, had to be suffi- 

 ciently flat to enable the lens to penetrate through its depth. 



Methods. 



As regards my own practice in making these cell-cultures, the 

 following note may be of use to other students ; though of course 

 different workers may operate differently. 



I first sterilise the plugged cells in the hot-air steriliser. When 

 these are nearly cool enough to han lie, I heat the quartz or glass 

 floor of the cell between two plates of talc (about 3x6 inches) held 

 in a large bunsen flame, and allow it to cool slightly. The cell is 

 then placed in position on its floor, and a small block of high- 

 melting paraffin cooled after sterilising is placed just outside the 

 cell. The temperature is still high enough to melt this, and the 

 liquid runs in by capillarity, and solidifies as the whole cools, 

 cementing the cell to its floor. 



When the culture is to be made I take such a prepared cell 

 several can be prepared and kept in sterile glass-covered dishes and 

 have ready the following : cover slips, a tube of infected gelatine (or 

 other medium), platinum, loops, sterile stiff gelatine, forceps, sterile 

 water, and any convenient rest, such as a small ring of brass. 



First, the necessary water is placed in the plugged cell, care being 

 taken to wet the cotton plugs and that some water shall be retained 

 on the inner side of each, whether a layer is spread over the floor or 

 not. 



Second, a cover-slip is sterilised between the talc-plates, and while it 

 is cooling the platinum loops are held in the flame and set aside to cool. 



Third, the cool sterile cover-slip is removed with the forceps, 

 and laid on the brass ring support. Then the drop is quickly put 

 on the centre of the slip, and the latter inverted on to the cell, so 

 that the drop hangs over the centre. 



* See ' Phil. Trans.,' vol. 183, 1892, B, p. 125. 



