68 



Journal of Applied Microscopy. 



improved by keeping it at a temperature 

 somewhat above the melting point for 

 several weeks. This is done readily by 

 placing it in large covered evaporating 

 dishes upon the steam radiators with 

 which our laboratories are heated. In 

 this way one can get a consistency at 

 least as good as that of old paraffin and 

 in a reasonable length of time. The 

 tendency to crystallize is diminished and 

 it is much less friable. There is no such 

 change in the melting point as in the 

 method of Graf Spee, and the results are 

 excellent. 



Frank Smith. 



University of Illinois. 



The Use of Soap for Imbedding 

 Plant Tissues. 



While engaged in a line of work 

 requiring the sectioning of the buds of 

 certain trees, it was found impracticable 

 to apply either the paraffin or collodion 

 method. The former method rendered 

 the objects much too brittle and both 

 methods required a longer time than 

 could conveniently be given. At the 

 suggestion of Professor Goodale, under 

 whose direction the work was being 

 done, experiments were made to deter- 

 mine what, if any, of the many soap 

 mixtures could be employed. The excel- 

 lent results obtained from the use of a 

 soap-mixture in this work have 

 prompted me to make known the details 

 in hopes many others may find it con- 

 venient to employ the same method. It 

 is especially adapted to work requiring 

 only paitial infiltration. 



Fleming, '73, used a soap containing 

 no glycerine, which he dissolved in 

 strong alcohol. One fatal objection to 

 this and all similar mixtures is that they 

 require too long to harden. Kadyi, 79, 

 gave a rather indefinite and complicated 

 method of making a soap solution by 

 dissolving in strong alcohol and then 

 adding enough water (to be determined 

 by repeated trial) to cause the solution 

 to form, upon hardening, a firm mass. 

 The uncertainty of the result by the use 

 of this method renders it unsatisfactory. 

 The details of this method can also be 

 found in the Vade-Mecum, Lee, '93. Sal- 

 ensky, '77, gives an account of a method 

 devised by Poelzam in which the soap 

 is to be cut into pieces and exposed to 

 the sun for several days before being 

 employed. None of the preceding 

 methods have proven at all satisfactory 

 in my experience. Pfitzer, '87, recom- 

 mended the use of a solution made by 

 dissolving at sixty degrees to seventy 

 degrees C as much glycerine soap as 

 possible in a mixture of equal parts of 



glycerine and ninety-five per cent, alco- 

 hol. This mixture, by cooling, hardens 

 in a very few minutes so that it is firm 

 enough to be sectioned. Poll, '89, gives 

 a similar method of preparing a gly- 

 cerine soap mixture. The latter two 

 were used by the authors on plant 

 tissues. 



My own experience has shown that 

 glycerine and alcohol together as the 

 solvent give the best results. The fol- 

 lowing suggestions I hope will make the 

 method perfectly definite and certain. 

 I have employed Pears' Transparent 

 Soap owing to the great uniformity of 

 this product and the resulting superior 

 transparency of the imbedding mass. 

 The soap is cut into small pieces and 

 dissolved in a mixture of equal parts of 

 ninety-five per cent, alcohol and gly- 

 cerine. For this purpose a temperature 

 of about 70 degrees C. has proved suffi- 

 cient, and the mixture while being made 

 should be kept in a flask provided with 

 a cotton plug to prevent too great evap- 

 oration. One gram of the soap should 

 be used for every cubic centimeter of 

 the alcohol-glycerine solvent. The result- 

 ing liquid is poured while warm into a 

 petri or other shallow glass dish and 

 there allowed to harden. This it will do 

 by cooling in five to ten minutes. The 

 cake thus obtained is cut into smaller 

 pieces, which can then be stored in a 

 bottle kept in a cool place. As this mass 

 will keep indefinitely, a quantity can be 

 made as directed and is then ready for 

 use at any time. 



Pieces of the prepared soap are placed 

 in a covered glass dish (a shallow sten- 

 der dish is very convenient) and the 

 mass remains liquid at forty degrees C. 

 The stender dish can be placed on the 

 ordinary paraffin oven. To secure thor- 

 ough penetration or infiltration it is well 

 to place the tissues in a dilute solution, 

 one that remains liquid at the tempera- 

 ture of the room for two to three hours, 

 and then into the melted soap mass, in 

 which the tissues should never be left 

 longer than five to ten minutes. A longer 

 stay in the warm solution is very apt to 

 injure delicate tissues and cause unde- 

 sirable swelling due to the alkaline char- 

 acter of the mixture. However, my expe- 

 rience has been that no difficulty in 

 staining will occur if one removes the 

 soap thoroughly with warm alcohol or 

 water. Final imbedding is best accom- 

 plished in a watch glass. The mixture 

 is very transparent and allows careful 

 orientation of the object. 



Sections are best cut free-hand with a 

 knife moistened with dilute alcohol. The 

 sections can, if desired, be fastened to 

 the slide with Mayer's albumen fixative. 

 I have employed successfully the follow- 

 ing stains: Delafleld's Haematoxylin, 



