The Culture Mount 26] 



Step 4. With a loop needle, a thill layer of the inoculated agar is spread 

 over the surface of the cover slips. 



Step 5. The organism is incubated at room temperature. When sulfieienl 

 growth has taken place, the cover slips may be removed with flamed 

 forceps. 



Step 6. For immediate study and temporary preparations, the cover slip 

 may be inverted on a drop of water on a slide and immediately examined. 



Step 7. For a permanent mount, the air bubbles arc removed by addition 

 of alcohol (95 per cent). The alcohol is drained off and a drop of lacto- 

 phenol is added with cotton blue. The cover slip is allowed to stand for 

 two minutes and then inverted on a microscope slide. After two or three 

 days, excess mounting medium is removed and the preparation is sealed 

 with asphalt varnish. 



For alternative methods to improve the seal (1) use the double cover 

 slip method, with the laetophenol cotton-blue mixture between cover slips 

 of dissimilar size, and after two to three days mount in clarite, which does 

 not turn yellow on standing; (2) allow the culture to dry, stain and mount 

 in clarite. 



2. CULTURE CHAMBER METHOD 



With this method, which is a modification of Henrici's culture chamber 

 method, the aerial portion of the colony may be studied independently of 

 the vegetative part. 



Step 1. A thin layer of dextrose agar is poured into a sterile Petri dish 

 and allowed to harden. 



Step 2. With a flamed hooked needle a hole is made through the agar, 

 the cut portion being discarded. The hole may vary in size or shape but 

 must be smaller than a cover slip. 



Step 3. The pathogenic fungus to be studied is inoculated at three or 

 four points on the rim of the hole. 



Step 4. A cover slip is placed over the hole, a small part being left un- 

 covered to admit an adequate supply of air. 



Step 5. Progress of the growth can be determined by viewing the growth 

 through the back of the Petri dish without opening the chamber. When the 

 culture is sufficiently developed, the lid of the Petri dish is removed. This 

 exposes the medium to outside contaminants, which may interfere with 

 prolonged study. 



3. WET INDIA INK PREPARATION 



Weidman and Freeman called attention to the value of a wet India ink 

 preparation in the differentiation of yeastlike growths. They advocated 



