624 Journal of Agricultural Research voi. xv, no. 12 



water after bleaching and kept in 70 per cent alcohol until they could be 

 stained. They were stained in safranin for one to two minutes, rinsed in 

 water, differentiated to the proper point with 50 per cent acid alcohol 

 and washed with fresh 50 per cent alcohol. They were then stained for 

 15 to 60 seconds in gentian-violet. The violet was followed at once by 

 the orange G with the slide held at an angle so that the orange G would 

 run off rapidly and carry the violet with it. Then a few drops of abso- 

 lute alcohol were mixed with the orange G which remained on the slide 

 to hasten the removal of excess violet, and the mixture v/as quickly 

 poured off. The mixing process and the pouring off of the mixture were 

 carried out as rapidly as it was possible to manipulate the reagents and 

 the slide. Then a few drops of fresh absolute alcohol were flowed quickly 

 over the slide. The absolute alcohol was replaced by clove oil, the clove 

 oil by xylol, and the mount then finished in balsam. The whole staining 

 and finishing process consumed only a few minutes and the results were 

 far superior to any obtained by following schedules calling for longer 

 staining. The success of the method depends on having good material 

 properly fixed, embedded, and sectioned, and on the use of fresh rea- 

 gents. The saving in time and the excellence of the finished slides more 

 than make up for the possible excess use of the more expensive alcohols 

 and oils. 



The method of using the iron-alum hematoxylin was also shortened by 

 cutting the time in the mordant by from 10 to 30 minutes, and the time 

 in the stain by from 5 to 30 minutes. The results were particularly good 

 for the study of the centrosomes and dividing nuclei. It is obvious that 

 the shorter methods outlined above have distinct advantages over the 

 longer methods generally advocated, provided they yield satisfactory 

 finished products. The short method ^ for the use of Flemming's triple 

 stain is recommended to investigators who have had trouble with this 

 more or less capricious combina;tion. 



MORPHOLOGY 



INFECTION OF PINE AND EXTERIOR INDICATIONS OF PRESENCE OF PARA- 

 SITE PREVIOUS TO SPORE FORMATION 



The actual process of the entrance of the sporidial germ tube in infect- 

 ing the pine has not been observed, and indeed the determination of the 

 actual point of natural infection is an exceedingly difficult problem. 

 Practically all infections first become evident by the etiolation or swel- 

 ling, or both, of the bark at the node or at some point in the internode. 

 The etiolation may be quite marked, as shown in Plate 48, A, represent- 

 ing a recent nodal infection. Internodal infections most often appear to 

 originate at the base of a leaf fascicle. A striking case of this type of 



» This short method, varied slightly according to the material to be stained, has been very successfully 

 used at the University of Wisconsin and Columbia University, and the author makes no claim for its 

 originality. 



