METHODS OF CULTURE AND OF EXAMINATION. 93 



(spiral or hook), whose capacity has been determined by 

 weighing. Especial importance is to be attached in this 

 connection to an equable distribution of the bacteria by 

 careful admixture. Of the mixture exactly the same care- 

 fully measured amount is introduced into ten cubic centi- 

 meters of gelatin, and then of this the same amount in a 

 third tube containing ten cubic centimeters, and so on. 

 After three, four, or five dilutions have been made, in 

 accordance with the approximate number of bacteria 

 originally present and the amount of inoculated mate- 

 rial, the gelatin is poured into plates, is permitted to 

 solidify, and the number of colonies that develop upon it 

 in the course of the next eight days is counted daily. 

 Generally the colonies are so dense upon the first and 

 perhaps also upon the second plate, that their enumeration, 



Fig. 24. Wolff hiigel's apparatus for counting colonies of bacteria upon plates. 



even with a lens or with a low power of the microscope, is 

 not possible. If the dilutions have been properly made, 

 there will always be one plate upon which the colonies can 

 be counted, and from this the numerical relations of the 

 colonies developed upon the other plates can readily be 

 estimated. The enumeration is greatly facilitated by the 

 use of a dark background divided into square centimeters, 

 and i or -i- of a square centimeter. Such an instrument is 

 the apparatus of Wolffhugel (Fig. 24), which consists 

 essentially of a glass plate divided into squares, and pro- 

 vided with a dark, dull background. 



In dealing with solid substances, I gram, j gram, etc., 

 is weighed, rubbed in a sterile mortar with from 5 to 10 

 cu. cm. of sterile bouillon or with physiologic salt-solution, 

 and the remaining steps are the same as those already 

 described. 



