230 Journal of Agricultural Research Voi. iv, N0.3 



methylene blue. This method gave nearly as good results as Guignard's 

 (6, p. 19), but no color differentiation was obtained. 



The Romanowsky simple method was not the most satisfactory, on 

 account of the lack of color differentiation, although it demonstrated 

 quite clearly the cell structures. 



Methylene blue, i to 1,000, in aqueous solution (flame-fixed), gave 

 good results; but, although repeated examinations were made of the 

 slides thus prepared, no red color was visible, the granules being indi- 

 cated only by a strong black-blue color. 



Romanowsky's compound stain (after Harris). No fixing v;as 

 done, since the methyl alcohol itself served as a fixing agent. Staining 

 was done by the mixture of methylene azure and methylene violet and 

 with eosin and methylene blue in methyl alcohol. This method, although 

 very good in showing the structure of the cells, failed in most cases to 

 give sharp differentiation. No real red color for chromatin w^as obtained, 

 as the one that would result from the staining of the haemoparasitic 

 protozoa. Some human blood stained by this solution gave the usual 

 color presentations. Leucocytes of all kinds, polymorphonuclear, macro- 

 nuclear, and eosinophylic, were seen, stained in their characteristic and 

 distinct colors. When A. chroococcum was stained by this method, the 

 network took a blue color and the interior of the meshes a pink one. 

 Repeated trials with the methylene-blue and glycerin method as used by 

 Mencl (i I ) failed to give satisfactory results; the cell always took the blue, 

 and the granules never took the red color that Mencl ascribes to them. 

 Vital stained cells present the same structure as the one described by 

 Mencl (PI. XXXI, fig. i). 



In the Romanowsky-compound method great attention must be paid 

 in diluting on the cover glass, since the solution tends to flow on the 

 underside and make a miscroscopical examination difficult. The time of 

 action for maximum differentiation varies with different objects. After 

 several trials the time limit for the action of the stain in these tests was 

 fixed at 10 minutes. After the addition of water to the stain on the 

 cover glass, the washing should be very rapid. The best preparations 

 were obtained when the washing was not prolonged over half a minute. 

 Distilled water was preferred in the washing. 



The cells of .4 . chroococcum stained differentially ; the network took on 

 a blue color, while the contents of the network meshes took on a pink 

 color. If the washing is prolonged, the pink color disappears and only 

 the blue remains in the cell, as in blood stained by this method. 



MICROCHEMICAL STUDIES 



Since the work of Mencl (11), Jones (9), Prazmowsky (15), and others 

 has shown that the structure of the cell of A. chroococcum is complex, 

 the present writer next proceeded to determine microchemically the 

 na*:ure of the different components. 



