34 



General Bacteriology. 



X 



Inasmuch as the first plate is invariably too thickly seeded to be of much service, 

 this gelatin tube is often replaced by a water blank, which is treated exactly as the gela- 

 tin tube No. 1, but is not of course "plated" but simply serves to dilute the material. 



REFERENCES. A. 124; H. 57; L. & K. 88; M. & W. 108; M. & R. 61; McF. 140; 

 171; P. 224; S. 72. 



SPECIAL DIRECTIONS. 



a. Make three gelatin plate cultures, as directed above, and inoculate with B. sub- 

 introducing a minute portion of agar culture (XIII) into tube No. 1, two loops of 



No. 1 into No. 2, and three of No. 2 into No. 3. Label, and when the gelatin has solidi- 

 fied, place plates in cool chamber (XV). 



b. Also make a "blank" plate from an uninoculated gelatin tube, observing all pre- 

 cautions to prevent contamination. This will serve as a control or check on your other 

 plates. If any colonies develop on this it indicates carelessness. 



EXERCISE XXVII. AQAR PLATE CULTURES. 



GENERAL DIRECTIONS. These are made in the same way as the gelatin plates ex- 

 cept that the high meltingpoint (96 C.) of agar makes it necessary to use boiling water 

 to melt it. Inasmuch as the vitality of vegetative bacteria is destroyed at a temperature 

 much above 42 C., it must be cooled down before inoculating, but as agar solidifies at 

 39-40 C. it must not, therefore, be cooled below that point. It is best to keep the melted 

 agar at about 42 C. for 10 minutes before it is inoculated. For this purpose a water- 

 bath should be so arranged that the temperature can be controlled 

 by means of a thermo-regulator. A cheap and yet satisfactory 

 03 W M ITl ^ arrangement is represented in Fig. 11. Inoculate, make dilu- 

 V\ 1 tions and pour as in case of gelatin, except that before the agar 



is poured, it is well to slightly warm the Petri dishes by placing 

 them iu the incubator at 38 C. for a few minutes, other- 

 wise the agar may solidify in lumps in the plate. In cooling, 

 agar shrinks somewhat, and in doing so water is expressed from 

 the solid jelly. In the incubator this condenses on the under 

 side of the cover of the Petri dish to such an extent that drops 

 run down on to the culture surface thus causing the developing 

 superficial colonies to "run." To obviate this the Petri dishes, 



FIG. 11. Water-bath for cooling 

 agar. 



when placed in the incubator, should be inverted. 

 REFERENCES. H. 61; L. & K. 94; M. & R. 66; N. 285; P. 225; P. B. C. 28. 

 SPECIAL DIRECTIONS, a. Make three agar plates of B. coli; use one loop of bou- 

 illon culture (XIII) for tube No. 1 and proceed as in XXVI. b. Place in incubator at 



EXERCISE XXV11I. ROLL-CULTURES (Esmarch). 



GENERAL DIRECTIONS. These are essentially plate cultures in which the medium 

 instead of being poured out into dishes is solidified in a thin, even layer on the inner surface 



of the test-tubes. This is best accomplished by means of a 

 piece of ice placed in a dish on a piece of cloth by which it 

 can be kept in the desired position (Fig. 12). A horizontal 

 groove is melted in the ice by means of a test-tube filled with 

 hot water. In this groove the test-tubes, inoculated as in case 

 cuUu '""of plate cultures, are rapidly whirled until the medium is thor- 



oughly set. Both agar and gelatin can be used, although gelatin cannot be used sue- 



FIG. 12. 



