COLLECTION, CULTIVATION, OBSERVATION 897 



washed thoroughly in running water for about 30 minutes. Rinse 

 them in distilled water. Transfer them through ascending series of 

 alcohol (50 to 95%). If counter-staining with eosin is desired, dip the 

 smears which were taken out from 70% alcohol, in 1% eosin in 95% 

 alcohol for a few seconds, and then in 95% plain alcohol. After two 

 passages through absolute alcohol and through xylol, the smears are 

 mounted one by one on a slide in a small drop of mixture of Canada 

 balsam and xylol. The finished preparations are placed in a drying 

 oven at about 60°C. for a few days. 



Other stains that are often used are as follows: 



Delafield's haematoxylin. If the stock solution is diluted to 1:5- 

 10, a slow, but progressive staining which requires no decolorization 

 may be made; but if stock solution is used, stain for 1-16 hours, and 

 decolorize in 0.5% HC1 water or alcohol. If mounted in a neutral 

 mounting medium, the staining remains true for a long time. 



Mayer's paracarmine. In slightly acidified 70% alcohol solution, 

 it is excellent for staining large Protozoa. If over-stained, decolorize 

 with 0.5% HC1 alcohol. 



Giemsa's stain. Shake the stock solution bottle well. By means of 

 a stopper-pipette dilute the stock with neutral distilled water (5-10 

 drops to 10 cc). Smears fixed in Schaudinn's fluid and washed in 

 neutral distilled water are stained in this solution for 10 minutes to 

 6 hours to overnight. Rinse them thoroughly in neutral distilled 

 water and transfer them through the following jars in order (about 

 5 minutes in each): (a) acetone alone; (b) acetone: xylol, 8:2; (c) 

 acetone: xylol, 5:5; (d) acetone : xylol, 2:8; (e) two changes of xylol. 

 The smears are now mounted in cedar wood oil (which is used for 

 immersion objectives) and the preparations should be allowed to dry 

 for a longer time than the balsam -mounted preparations. 



Feulgen's nucleal reaction. The following solutions are needed. 



(a) HC1 solution. This is prepared by mixing 82.5 cc. of HC1 (spe- 

 cific gravity 1.19) and 1000 cc. of distilled water. 



(b) Fuchsin-sodium bisulphite. Dissolve 1 gm. of powdered fuchsin 

 (basic fuchsin, diamant fuchsin or parafuchsin) in 200 cc. of distilled 

 water which has been brought to boiling point. After frequent shak- 

 ing for about 5 minutes, filter the solution when cooled down to 50°C. 

 into a bottle and add 20 cc. HC1 solution. Cool the solution further 

 down to about 25°C. and add 1 gm. of anhydrous sodium bisulphite. 

 Apply stopper tightly. Decolorization of the solution will be com- 

 pleted in a few hours, but keep the bottle in a dark place for at least 

 24 hours before using it. 



(c) Sulphurous water: 



