704 Botanical Microtechnique 



immersing in the destaining agent for 1 to 2 min., rinse in distilled 

 water, examine with the microscope, and repeat the brief immersion 

 in ahrni until the nuclei and pyrenoids are sharply differentiated. 

 After the last rinsing in distilled water, wash thoroughly in tap water. 

 Dehydration and mounting may be accomplished by one of the several 

 methods outlined below. 



THE VENETIAN TURPENTINE METHOD 



The Venetian tmpentine method fields excellent preparations, 

 but this method is likely to be supplanted by modern methods using 

 a variety of synthetic organic solvents and synthetic resins. Therefore, 

 only a brief otitline of the method will be given, and the interested 

 reader is referred to Chamberlain for details (1932) . Kill and stain 

 the material, and dehydrate by the glycerin evaporation method. 

 Rinse otit the glycerin in 95% alcohol, then complete the dehydration 

 in at least three changes of absolute alcohol. Transfer to 10% Vene- 

 tian turpentine in absolute alcohol. Eliminate the alcohol by evapora- 

 tion. Wlicn the material is in thickened turpentine, mount the de- 

 sired amount of the material in a drop of that medium. Dry the 

 slides in a horizontal position. Preparations made by this method are 

 durable and the stains arc permanent. 



THE BUTYL ALCOHOL-RESIN METHOD 



1 he term resin is used here in the broad sense to include Canada 

 balsam as well as the increasing ninnber of synthetic resins. Test the 

 solubility of the currcnth available mounting resins in (niJixdyoiis 

 normal and tertiary butyl alcohol and in dioxan. and use the solvent- 

 resin combination that has the highest transparency and least color. 

 The following tertiary biuyl alcohol - Canada balsam j^rocedure 

 serves as an example of the series of operations in a whole-moiuit 

 method. Stain and wash as before. Transfer a small quant it\ of the 

 material into a watch glass of 50%, alcohol, and obser\e with the 

 microscope. If there is nuich distortion, try 20% alcohol on another 

 batch. Well-hardened material can Avithstand 50',. W'luii tiu' proj^er 

 starting ]X)int lor dehydration is established, carrv tlic material iu 

 steps of 20 to approximately 70% alcohol. Add to the 70% a few 

 drops of sto(k solutiou ol (ouiucistaiii — eosiu ^. erythrosin B, or 

 fast green, saturated solution in al)s<)liue akohol or in melhvl Cello- 

 solve. Lea\e in the coiuuerstain until slightly o\erstained. This may 

 recpiire 1 to 12 hi. Rinse in 70%, ahohol. and transfer thiough the 

 r()ll()\vin!; sii ies at 1/2- to l-hr. inter^■als: 



