2 66 PREPARATION OF SECTIONS 



time ratios in the stains will need to be varied for different 

 materials. Iodine green may be used in place of the cyanin. 

 It is cheaper than cyanin and is easier to work with. 



Cyanin and erythrosin can be used for sections cut free- 

 hand or in any other way, and loose sections may be stained 

 in watch glasses. A beautiful differentiation of the protoplast 

 in shades of gray is obtained by iron alum-hgematoxylin. 

 Place the sections for two hours in a 3 per cent, aqueous solu- 

 tion of ammonia sulphate of iron, then wash in water for half 

 an hour and place for about ten hours in a 0.5 per cent, aqueous 

 solution of hasmatoxylin that has ripened in a bottle plugged 

 with cotton, to let in the air, for two months. Remove from 

 the haematoxylin, rinse in water five minutes, and place again 

 in the iron alum to reduce the too intense stain. Keep watch 

 of the bleaching process under the microscope until the parts 

 of the protoplast appear no longer muddy, but still well defined. 

 Now wash in water for an hour or more, and pass through 

 alcohol and xylene, and mount in balsam. After the last wash- 

 ing in water the sections may, if desired, be very lightly counter- 

 stained in a weak aqueous solution of fuchsin or orange G, 

 and then, after again rinsing, be carried through alcohol and 

 xylene for mounting in balsam. 



It must be borne in mind that always when sections in water or 

 aqueous stains are to be mounted in balsam they must pass from 

 the water into 95 per cent, or absolute alcohol for dehydration, 

 and then into xylene which is a solvent of balsam. If it is 

 found that the sections look milky or opaque when taken from 

 the alcohol to xylene it is a sign that dehydration was not com- 

 plete, the alcohol was not strong enough or the sections were 

 left in it for too short a time. Opaque sections of this kind 

 will clear up more or less after long standing in xylene. 



Imbedding in Celloidin. — Sometimes material that is not 

 suitable for sectioning free-hand will also not give good results 

 when imbedded in paraffin, on account of its size, hardness, or 

 brittleness. In such cases we may get help in celloidin or 

 collodion (gun cotton) for imbedding. The ])rocess of obtain- 



