100 



LABORATORY EXERCISES IN BACTERIOLOGY. 



,1 



The serum as it passes into the tubes is usually more or less red and somewhat 

 turbid from the admixture of corpuscular elements of the blood. The tubes are there- 

 fore allowed to stand for several hours longer in the refrigerator, when it will be found 

 that the red blood will have settled to the bottom and the supernatant, clear, straw- 

 colored serum can with little difficulty be forced from each tube disconnected from 

 the series, as from a wash-bottle, by placing a plug of sterile cotton in the open end 

 of the short tube and blowing through it. When this is done, the long end of the 

 other tube should have been slightly withdrawn from the red sediment in the bottom 

 of the sedimentation tube lest this be forced into the culture tubes (Fig. 30). 



Other methods of distribution are often suggested or may be devised as circum- 

 stances demand. One of the most simple and efficient is accomplished by means of a 

 sterilized pipette (of fifty or one hundred cubic centimeters capacity) having the 

 upper end protected from the entrance of organisms by a sterile cotton plug, the serum 



being drawn into such a pipette by suction by 

 the mouth of the operator. 



After distribution of the serum to the cuK 

 ture tubes (or dishes, if desired) the further 

 manipulation consists in subjecting it to low 

 heating (Pasteurization 60 C.) for an hour 

 each day for five or six days, in order to insure 

 the medium from the influences of organisms 

 which have possibly gained entrance to it 

 during the steps of collection or distribution. 

 Upon the sixth or seventh day, with the tubes 

 adjusted in a slanting position to procure a 

 large surface of exposure, the serum is gradu- 

 ally heated above 70 C. until it assumes the 

 appearance of a stiff, clear, straw-colored jelly. 

 During this stage the material must be fre- 

 quently examined lest it be overheated. Differ- 

 ent specimens coagulate at temperatures vary- 

 ing from 70 to 85 C. and from half an hour 

 to two hours' exposure. Usually, the longer 

 periods of exposure and lower temperatures 

 applied are followed by the most sightly pro- 

 duct. Before the medium is employed for any 

 important work, one or two tubes should be 



placed in the incubator at body temperature for the development of any organisms 

 which may possibly be present. Should growth take place in these tubes, the entire 

 lot of tubes are to be doubted and may be either rejected or tried in the incubator, when 

 the sterile ones are selected ; or all may be converted into the opaque white serum by 

 heating them in the oven to 90 C. for about ten or fifteen minutes, after which they 

 may with safety be sterilized in steam by the fractional method. In Pasteurizing 

 and coagulating the serum in the above manner it is convenient to employ a special 

 warming chamber, known as a serum inspissator, consisting of a chamber surrounded 

 with a water-bath to preserve evenness and uniformity of temperature, the apparatus 

 being provided with legs which may be adjusted so as to allow the tubes in the interior 

 to lie in a slanting position. In such a chamber the tubes should be placed upon a 

 layer of cotton or other material to keep them from direct contact with the walls, 



FIG. 30. DISTRIBUTION FROM SEDI- 

 MENTATION TUBE TO CULTURE 

 TUBES ; TUBULE a RAISED ABOVE 

 RED SEDIMENT IN SEDIMENTATION 

 TUBE. 



