Alga 



WHOLEMOUNTS IN RESINOUS MEDIA 



65 



and takes its name, as do so many other 

 resins, from the place from whicli it was 

 first exported to England. The commonest 

 commercial use of this material today is in 

 the preparation of artist's pigments, so 

 that it is usually better secured from an 

 artists', than from a scientific, supply 

 house. Man}' substitutes and impure speci- 

 mens are on the market and, as the only 

 value of Venice turpentine lies in its per- 

 fect miscibility with alcohol, any specimen 

 should be tested b}^ seeing whether an 

 equal volume of tlie balsam and of 95% 

 alcohol will make a perfectly clear mix- 

 ture. If they do not, the specimen is not 

 true Venice turpentine and is worthless 

 for the technique which follows. 



Zimmerman 1896, p. 18, recommends 

 that, in any case, the raw resin should be 

 diluted with twice its own volume of 95% 

 alcohol, filtered, and then heated until the 

 alcohol is evaporated from it. Tliis method 

 is, however, dangerous, unless the atmos- 

 pheric humidity is practically nil. It is 

 much simpler to get a first-class specimen 

 of Venice turpentine in the first place. 



This medium was first recommended for 

 the preparation of botanical wholemounts 

 by Pfeiffer and Wellheim 1892 (23632, 

 8:29) but did not come into general use 

 until it was reintroduced by Chamberlain 

 1915 (p. 97). This writer, however, com- 

 plains that the original directions of Pfeif- 

 fer and Wellheim were diffuse; he cites, 

 not their original paper on the mounting 

 of objects in Venice turpentine, but an- 

 other paper on the preparation of fresh- 

 water algae (Pfeiffer and Wellheim 1894; 

 10606, 26:674). The present description is 

 drawn from all the sources cited. 



We will assume that we are dealing with 

 Spirogyra, a form notoriously difficult to 

 prepare as a satisfactory wholemount. It 

 may be collected in quantity almost any- 

 where in the world, and should be trans- 

 ferred immediately after collection to a 

 large volume of any chromic-acetic fixa- 

 tive; the formula of Lavdowsky 1894 

 (Chapter 18, F 6000.0010) is excellent for 

 the purpose. After the masses of algae 

 have been in the solution for a day or two, 

 they should be washed in running water 

 for 24 hours and then pieces should bo 

 selected for mounting. These pieces should 



be relatively straight, and about y> inch 

 long, for it is a waste of time to take great 

 masses of algae through the compUcated 

 l)rocesses which follow, and then to select 

 finally only the few pieces which one de- 

 sires to mount. The selected pieces should 

 be carefully passed through 15%, 30%, 

 50% and 70% alcohol and finally into 

 90% alcohol in which they are to be 

 stained. 



Some specimens are so delicate that 

 they will not stand transference from 

 water to alcohol, no matter how gradual 

 the transition phases may be, and for 

 these the method of Chamberlain may be 

 adopted. The algae are transferred from 

 water to 10% glycerol antl the water then 

 evapoiated until they are in pure glycerol. 

 The glycerol is then washed out with 95% 

 alcohol without risk of tlie specimen col- 

 lapsing. This washing must, however, be 

 thorough. 



Assuming that we now have the speci- 

 mens in 95% alcohol, it is recommended 

 that they be stained by the technique of 

 Chamberlain (Chapter 20, DS 13.5 Cham- 

 berlain 1915). For this there will be re- 

 quired a 1% solution of phloxine in 95% 

 alcohol, a similar strength solution of ani- 

 lin blue in the same solvent, and a 0.1% 

 dilution of hydrochloric acid, also in 95% 

 alcohol. The specimens are transferred 

 from alcohol to the 1 % phloxine solution , 

 where they remain for about 24 hours. 

 They are then rinsed in alcohol for a min- 

 ute or two, or until most of the excess has 

 been removed, and transferred to the ani- 

 lin blue solution. They should remain in 

 this until sufficient blue color is showing 

 in the cytoplasm and until the cell walls 

 themselves have just started to take this 

 blue. They should not, however, remain 

 in the blue for sufficiently long to obscure 

 the bright red color of the nuclei. Cham- 

 berlain suggests that from three to thirty 

 minutes may be necessaiy, but the writer 

 has never had to use a longer period than 

 five. It is obviously desirable to experi- 

 ment witli a few filaments until one has 

 esta])lished the correct time and then 

 stain all the rest of the filaments together. 

 After the filaments have been stained in 

 the blue, they should be transferred to the 

 acid alcohol where they should remain 



