A NOTE ON MATERIAL AND METHODS 7 



method* is very good for all amoebae, but usually not reliable — owing 

 to unequal penetration — for staining cysts. 



I have obtained beautiful preparations of amoebae and their cysts, 

 and of sections of intestinal ulcers and other tissues containing E. histo- 

 lytica, by employing a modified form of Mann's stain. As I have used 

 this method for some years, and taught it to many people, I may mention 

 it here. I use Mann's well-known methylblue-eosin mixturef made up 

 in the usual way, but differentiate the preparations with a very dilute 

 solution of Orange G in 70 per cent, alcohol — instead of using the 

 alcoholic solution of caustic soda which he employs. Very fine results 

 can be obtained by this method — not only with tissues and amoebae, but 

 often with cysts as well. For sections I have also found Borrel's magenta 

 and picro-indigo-carmine method very useful : but a modification of 

 this, in which I use acid fuchsin instead of basic fuchsin, gives even 

 better results with sections containing E. histolytica, as the amoebae can 

 be differentially stained by this method. Safranin may also be used 

 instead of magenta, and gives excellent results. For demonstrating 

 glycogen in the cysts of amoebae, Best's specific carmine stain:]: is very 

 useful — both as a control for the reactions obtained with iodine, and 

 as a method by which permanent preparations showing glycogen can 

 be obtained. Cysts may be fixed previously in Schaudinn's sublimate- 

 alcohol solution, or with Carnoy'h chloroform-alcohol-acetic-acid mix- 

 ture : and if it is desired to show the nuclei as well as the glycogen, 

 they can be coloured by previously staining the cysts in Weigert's 

 alcoholic iron-chloride haematoxylin — without removing the glycogen 

 or affecting its staining powers. 



All the specimens figured in the Plates have been drawn, with the 

 aid of the camera lucida, at a uniform and exact magnification of 

 2500 diameters. All the drawings were made from preparations examined 

 under a 2 mm. apochromatic objective (Leitz), with N.A. = 1.40, using 

 compensating oculars and an achromatic aplanatic condenser. The 

 methods by which the specimens figured were fixed and stained are 

 all noted in the descriptions of the Plates. I will only add here that 

 for studying and drawing organisms stained with red stains — such as 

 fuchsin or carmine — I have found it an advantage to use a green light 

 for illumination, as details can then be resolved with greater ease and 

 precision. I have found a Wratten colour screen (" B. Filter," No. 58), 

 placed below the condenser of the microscope, very useful for this 

 purpose. This is, of course, a general method, and not one peculiarly 

 suited to the study of amoebae. 1 mention it here merely because all 

 specimens actually stained red, but depicted in black in Plates Ill-V, 

 have been drawn from preparations examined in this manner. 



• Vide Dobell (1914). 



t Vide G. Mann : " Physiological Histology " (1902), p. 216. 



; Vide F. Best (1906) : Zeitschr. f. wiss. Mikrosk., xxiii, 319. 



