72 



THE ART OF MAKING MICROSCOPE SLIDES 



Staining 



about three or four minutes in the case of 

 a thin smear, or for five to ten minutes in 

 the case of a thick one. It is then trans- 

 ferred to distilled water to await staining. 

 It occasionally happens that a slide 

 must be fixed in one of the conventional 

 fluid fixatives. This is done with the same 

 petri-dish and glass-rod setup as is used 

 for vapor fixation, but in this instance the 

 fixative is carefully poured into the petri 

 dish, which must be level, until it has 

 reached such a depth that, when the slide 

 is laid across the glass rods, the under side 

 of the slide with the smear on it is in con- 

 tact with the fluid while the upper part is 

 free from fluid. If the smear is reasonably 

 tliin and is laid carefully in place, it usu- 

 ally will not become detached. Thick 

 smears, particularly those made with 

 fluids containing very little protein, will 

 not stand this treatment; they must either 

 be fixed in the vapor phase, or else the 

 fluid itself must be mixed with a small 

 quantity of an adhesive, such as Mayer's 

 albumen (Chapter 28, V 21.1 Mayer 

 1884). 



Staining Smears 



Blood smears are so universally stained 

 with one or another of the methylene blue- 

 eosinate mixtures (Chapter 20, DS 13.1 

 and 13.2) that it comes as something of a 

 surprise to most people to learn that any 

 stain which is suitable for sections may 



also be employed for smears. The ad- 

 vantage of methylene blue-eosinates for 

 blood films is that the solvent methanol 

 acts as a fixative so that they are stained 

 and fixed in the same operation. When a 

 blood smear is to be used for diagnostic 

 purposes, these techniques are excellent, 

 because the appearance of the various 

 types of white corpuscle under this treat- 

 ment is known to every technician. For 

 research studies on the blood, however, it 

 is strongly recommended that the worker 

 experiment, first by fixing the blood film 

 in osmic vapor in the manner described, 

 and secondly by applying to it some other 

 of the complex techniques described in 

 Chapter 20. For materials other than 

 blood there is no limit to the type of stain- 

 ing which may be employed, though it 

 must be remembered that very thin films 

 require a stain of considerable intensity if 

 the finer structures are to be made out. 

 Thus, for example, a thin smear of mam- 

 mahan spermatozoa is best stained by one 

 of the very dense iron hematoxylin tech- 

 niques such as that of Biitschli (Chapter 

 20, DS 11.111 Butschh 1892). The method 

 for the application of these stains to 

 smears differs very little, in most cases, 

 from the method for the application of the 

 same stains to slides, and no specific in- 

 struction need be given. Bacteriological 

 staining methods, which differ from those 

 used in botany and zoology, are given in 

 Chapter 23 under the heading DS 23.2. 



Typical Example 



Demonstration of Monocystis from the Seminal Vesicle of an Earthworm 



Few sporozoans are available for class 

 demonstration purposes and the choice is 

 practically limited to the inhabitants of 

 the intestines of a cockroach or to the 

 specimen at present under discussion, 

 Monocystis. 



The advantage of Monocystis is that all 

 the forms from the sporozoite to the 

 trophozoite occur in the seminal vesicle of 

 the earthworm, and may therefore be 

 made available on a single smear. The 

 degree of infection among earthworms 

 varies greatly, but it has been the author's 

 experience that the larger the earthworm 

 the more hkely the chance of a heavy in- 



festation. But it is no use making a whole 

 lot of smears for class demonstration pur- 

 poses until one has satisfied oneself by a 

 preliminary survey of a single smear that 

 the material will be satisfactory. 



There is no need to kill or anesthetize 

 the earthworm, which' is simply pinned 

 down in a dissecting tray and sUt from the 

 anterior end to about the 16th or 17th seg- 

 ment. The edges of this slit are pulled 

 back and pinned into place disclosing the 

 large white seminal vesicles. 



There should be available, before mak- 

 ing the smear, a petri dish in which are a 

 couple of short lengths of glass rod, a sup- 



