114 SPECIFIC EXAMPLES OF SLIDE-MAKING 



be thrown away. Those slides which are perfectly adherent are warmed over a 

 flame until the wax is melted and then dropped into a jar of xylene, where 

 they remain until a casual inspection shows the whole of the paraffin to have 

 been removed. They are put into another jar of xylene for at least five minutes 

 and then into a jar of equal parts xylene and alcohol for a further five min- 

 utes. This treatment is followed by five minutes in absolute alcohol and then 

 by direct transference to distilled water. After the slides have been in distilled 

 water for a few minutes, each slide should be lifted and inspected to make 

 sure that the water is flowing uniformly over both the slide and section. If it 

 tends to be repelled by the section or a meniscus is formed around the section, 

 this is evidence that the wax has not been sufficiently removed, and the slide 

 must be transferred first to 96 per cent alcohol to remove the excess water, 

 then to absolute alcohol until perfectly dehydrated, and finally through abso- 

 lute alcohol to xylene, where it remains until the wax has been completely 

 removed before being brought down again as previously indicated. The slides 

 may be taken down through xylene and alcohol one at a time and accumulated 

 in distilled water until they are required. When all slides have been accumu- 

 lated in distilled water, they are transferred to the celestin blue B staining solu- 

 tion. The time in this varies, but 10 to 15 minutes will probably be sufficient 

 to stain the nuclei black. One of the most useful features of this stain is that 

 it is almost impossible to overstain in it. It is also possible to leave sections 

 overnight without staining the cytoplasm to a degree which requires diffisren- 

 tiation. After the nuclei are black, therefore, or after a time convenient to the 

 operator has elapsed, the sections are transferred to fresh distilled water where 

 they are thoroughly washed. Then each slide is taken individually and dipped 

 up and down in the eosin solution until a casual inspection shows the back- 

 ground to be yellowish pink. The staining of the background in a case like 

 this is entirely a matter of choice; some people prefer a faint stain and others 

 a darker stain, although it must be remembered in judging the color that the sec- 

 tion will seem darker after it has been cleared than it does in water. 



As soon as the time required to produce the desired degree of staining has 

 been found by staining one slide, the remainder of the slides are placed in the 

 eosin solution, left the appropriate time, and then transferred and left in dis- 

 tilled water until no more color comes away. The slides are passed from distilled 

 water to 96 per cent alcohol where they are left for about five minutes. Then they 

 are passed to fresh 96 per cent alcohol where they are left for five or six min- 

 utes before being passed to absolute alcohol. The purpose of using the 96 per 

 cent alcohol is not to diminish diffusion currents but simply to save diluting 

 the absolute alcohol by passing slides directly from water to it. After the slides 



