Journal of Applied Microscopy. 227 



2. Add enough chloroform to make a two-thirds solution, and let it remain 

 from four to eight hours. 



3. Transfer to pure chloroform and leave from six to twelve hours. 



5.— IMBEDDING IN PARAFFIN. 



The objects are now ready for the paraffin. This should be of good quality, 

 with the melting point at 49 degrees or 50 degrees C. The paraffin must be added 

 gradually, in the following manner : add small pieces of cold paraffin to the 

 chloroform in which the objects are, sufficient to form a cold saturated solution. 

 After the cold chloroform has taken up all the paraftin possible, say after about 

 six or eight hours, the objects must be gradually brought into the hot water oven. 

 This may be of various designs and sizes. A square oven with a side door is 

 very convenient and cheap. The oven should be kept at a uniform temperature 

 of about 52 degrees C. The bottle may first be placed on top of the oven, and 

 then inside with the door a little open, and finally with the door closed. When 

 warmed up to the temperature of the oven, melted paraffin, kept in a suitable 

 dish in the oven, may be added from time to time, at intervals of two or three 

 hours. At the same time some of the mixture of chloroform and paraffin is poured 

 off until the objects are in pure melted paraffin, with all traces of chloroform re- 

 moved. The objects should stay in the oven at least a day, and several days 

 will do no harm if the temperature is uniform. I usually take two days for the 

 operation. One day, however, is usually long enough unless the objects are very 

 large and difficult to penetrate. 



6.— MAKING THE CAKE. 



The final imbedding can be easily done in the following manner : use a Petri 

 dish of proper size, 80, 120, or 150 mm. in diameter, depending on the amount 

 of material to be imbedded ; or the paraffin imbedding dish described by the 

 writer in Vol. 1, page 11, of this journal. Before imbedding, apply a very thin 

 coat of a 50 per cent, aqueous solution of glycerine to the parts of the dish with 

 which the paraffin will come in contact, and pour in a suitable amount of melted 

 paraffin to make the cake. The objects being in the bottle with the cork, turn 

 the bottle upside down and allow the objects to settle on the cork. Then 

 remove the cork and let the paraffin in the bottle, with the objects, fall into the 

 dish. The objects may be arranged in the paraffin with hot needles. Put the 

 dish quickly into cold water, but do not let the water flow into the dish until the 

 paraffin is hard enough to bear the weight of the water without being distorted. 

 The paraffin cake must be cooled very rapidly, and this is usually done best in cold 

 flowing water. After the cake is thoroughly hardened it is carefully removed 

 from the dish and laid aside until used. When the objects are once properlv 

 imbedded they can be preserved for an indefinite period if kept in a cool place. 

 The bottle in which the objects were kept while passing through the parafiin may 

 be used for the same purpose for subsequent imbeddings. After the objects are 

 in pure chloroform they can be poured into this bottle, which will already have 

 some paraffin adhering to its walls. 



John H. Schaffner. 



Ohio State University. [To be Continued.) 



