[Vol. 6 

 208 ANNALS OF THE MISSOURI BOTANICAL GARDEN 



the calculated value, the greatest divergence being .2. In the 

 alkaline range the concentration of the OH ions in the last solu- 

 tion was beyond the range of the indicator, so it has been desig- 

 nated as 10.0+. The ten solutions, termed a series, are as 

 follows: P H 2.8, 3.1, 4.4, 5.0, 6.2, 7.0, 7.4, 8.8, 9.6, and 10.0+. 



Small portions of each of the solutions were transferred to 

 sterile test-tubes permanently labeled and fitted with rubber 

 stoppers, through each of which passed a glass rod drawn to a 

 blunt point. All transfers of a solution were made with its par- 

 ticular rod, thus avoiding all chances of mixing solutions. 

 Fresh solutions were placed in the tubes from time to time, and 

 verifications of hydrogen ion concentration frequently made. 



Method of culture. — The method of culture was based primar- 

 ily on the hanging-drop or Van Tieghem cell. The glass cylinders 

 employed were perfectly ground at each end and measured 18.0 

 mm. in diameter and 9.0 mm. in height, possessing therefore a 



volume of 2.3 cc. The cylinders were cemented to the slide by 

 means of wax; the tops of the cylinders were then coated with a 

 thin ring of vaseline, and the cells completed by sealing cover 

 glasses to the tops of the cylinders. A small nick was made in 

 each ring of vaseline, prior to sealing, so that equilibrium of air 

 pressure might exist when the cultures were placed in the incu- 

 bator. About 15 minutes later, the cultures were examined and 

 the cover glasses slightly pressed to the cylinders in order to 

 insure a perfect sealing. Two cells were placed on each slide 

 and labeled by gumming numbered and lettered labels to the 

 center of the slip. 



Four or five drops of the same solution as that to be used in 

 the culture were placed in the bottom of the cell, the object 

 being to establish a complete equilibrium of vapor pressures in 

 the cells and to prevent changes in the concentration of the 

 solution tested, as shown by Clark ('99). A few drops of the 

 same solution were also placed on a sterile slide, and spores 

 transferred from a pure culture to the slide by means of a 

 sterile platinum needle. A solution of spores was thus made 

 and thoroughly stirred in order to prevent the spores from 

 adhering in bunches. A drop of the spore solution was trans- 

 ferred from the slide to the cover glass by means of a clean, 





