XIII, D, 5 Haughwout: Protozoa of Manila and Vicinity: I 185 



A good general stain, which often gives an excellent cyto- 

 plasmic effect in addition to the nuclear stain, is the formula 

 of Delafield, which is compounded thus: 



Hematoxylin crystals (grams) 4 



Ethyl alcohol, 95 per cent (cc.) 25 



Saturated aqueous solution of ammonia alum (cc.) 400 



Dissolve the hsematoxylin in the alcohol and add it to the 

 alum solution. Allow the mixture to ripen in the light in a 

 bottle lightly stoppered with cotton. At the end of three or 

 four days filter the solution and add: 



Glycerin (cc.) 100 



Methyl alcohol (cc.) 100 



This solution should be allowed to ripen further for four or 

 five weeks before using. It keeps well, but has a tendency to 

 redden in the course of time. When this occurs, add a small 

 amount of 1 per cent alum solution or a crystal or two of alum. 

 The stain is best used in dilute solution, for it is very powerful 

 and penetrating. 



A useful formula is one that I have used extensively for many 

 years as have several of my colleagues. This stain should be 

 made up as follows: 



Concentrated Delafield's heematoxylin (cc.) 10 



Distilled water (cc.) 90 



Glacial acetic acid (cc.) 0.5 to 1 



Chloral hydrate crystal (gram) 0.5 



Staining may be done with this mixture by either the pro- 

 gressive or the regressive method. It often gives a good flagel- 

 lum stain by the progressive method. I have obtained some 

 fine preparations of the malarial parasite by prolonged staining 

 in the mixture. 



The carmine stains often give brilliant results, especially if 

 used after mercury fixation. Simple borax carmine or the picro- 

 carmine of Hoyer, Ranvier, or Weigert are to be recommended, 

 although I have secured my best results with Hoyer's formula. 

 When staining by the regressive method with any of the hsema- 

 toxylin or carmine stains named, differentiation should be carried 

 out in 70 per cent alcohol very slightly acidified with hydro- 

 chloric acid. 



This, of course, does not exhaust the list of methods that may 

 be employed with the Protozoa. For others the reader must 

 consult the general and special works on the Protozoa. 



When protozoa are very abundant in a culture their collection 

 and staining is a relatively simple matter. Under such conditions 



