334 METHODS IN PLANT HISTOLOGY 



late cone. Trim away most of the bud scales and cut a slab from oppo- 

 site sides, leaving a piece 2 or 3 mm. thick to be imbedded. The bud, 

 and also selected pieces of the young stem, will show the structure of 

 the young leaf. 



Toward the end of the season, the first-year shoot is rigid enough to 

 be cut without imbedding. This shoot and also two-, three-, and four- 

 year shoots are generally cut without imbedding. When living mate- 

 rial is cut in this way, the sections should be placed in water as they 

 are cut, and then fixed in formalin-acetic acid for 24 hours. Run them 

 up through close grades of alcohol to 85 per cent, where they should 

 be left for 24 hours. Then, after | hour in 70 and ^ hour in 50 per cent 

 alcohol, they can be stained. With this care, which is seldom taken, 

 the protoplasm should not pull away from the cell wahs of the cortex. 



It is quite possible to cut paraffin sections of two-, three-, and four- 

 year pine stems. Cut into pieces not more than 1 cm. long; fix in for- 

 malin-acetic-alcohol for several days — or months; wash in 70 per cent 

 alcohol, 8 hours; 85 per cent alcohol, overnight; 95 per cent, overnight; 

 100 per cent, overnight. Make the transfer to xylol very gradual. 

 Place the pieces of stem in absolute alcohol at one side of a large Petri 

 dish and put a few drops of a mixture of equal parts of xylol and 

 absolute alcohol into the dish at the side opposite the material. Every 

 half-hour or so, put in several drops of the alcohol-xylol mixture. 

 When the quantity of liquid has nearly doubled, pour off about half 

 of it, and continue the dropping. Then drop with pure xylol. When 

 the material looks as if it were nearly cleared, transfer to pure xylol. 

 Pieces of stem 1 cm. in diameter and 5 mm. thick should be cleared in 

 about 24-36 hours. Then put some paraffin into the xylol, about 3 

 hours at room temperature; put in more and leave it on the bath for 

 3 hours; then inside the bath, 30-36 hours, with three changes of 

 paraffin. Imbed. The time for mixtures of paraffin and xylol is the 

 minimum. 



For the structure of the adult stem, select a clear board (Pinus 

 strohus is very good) ; and for transverse sections, cut out pieces 2 cm. 

 long and trimmed to give sections 5X8 mm. ; for longitudinal sections, 

 use pieces 2 cm. long, with 5 and 10 mm. for the other two faces. 



Cut from the face which will give sections 5X10 mm. Orient care- 

 fully, so that the longitudinal radial sections shall be exactly parallel 

 with the rays, and the longitudinal tangential sections exactly tan- 

 gential to the rays. 



