THE MYXOSPORIDIA, OR PSOROSPERMS OF FISHES. 119 



VII.— MICROSCOPIC TECHNIQUE. 



The older observers used no reagents beyond acetie aeid, potassium 

 hydrate, etc. Biitschli^ was the first to use a staining reagent. He 

 believed that alum carmine stained nuclei in the ectoiDlasm. The 

 first observer to emjiloy modern technique was Ilenneguy.^ Subse- 

 quently Thelohan^ employed similar technique, and Pfeiffer'* devotes 

 some space to the technique of protozoan investigation. Finally Hen- 

 neguy and Thelolian ^ give a few additional remarks upon this subject. 



The following is a summary of the methods recommended: Fixing 

 and hardening preferably by chromic or osmic acid or both (Perenyi's 

 or Flemming's liquids ''') or corrosive sublimate solution. Washing out, 

 dehydration, parafitining, sectioning as usual. Affixing to the slide by 

 Mayer's albumen. Where alcohol-fixed material is the only kind avail- 

 able, much may be gotten out of it in the way of study of the spore. 



Dissociation (1 per cent osmic acid solution ; Ripart and Petit's liquid) 

 shows certain facts better than the section method. 



Sections are necessary to determine the seat, and, above all, to 

 follow tlie different stages of development. 



Culture in the blood (overhanging drop method) is recommended by 

 Pfeifter for the study of development. 



Stains:" For alcoholic specimens, carmine; above all other forms 

 hydrochloric acid alcohol carmine is very reliable. For chrom-osmium 

 (and may be tried on alcoholic) specimens, especially gentian violet, 

 double stain with the violet by eosin. Safranin, by Henneguy's 

 method,** evinces an electivity valuable in the study of development 

 where we have to do with the most comi)lex phenomena of cellular life 

 under circumstances in which the small size of the elements renders 

 observation extremely difiicult. The sections must be decolorized in 

 clove oil for a very long time. Small stellate-grouped masses of crys- 

 tals, which are often precipitated and whose presence is very annoying 

 in the subsequent study of the section, may be easily removed by suc- 

 cessive alternate washings of the latter in chloroform and bergamot oil. 



Valve separation : Most certainly efiected by sulpliuric acid (cold, 

 concentrated). 



Vacuole : Best shown by very dilute iodine water (with potassium 

 iodide). 



' Ztschr. f. wiss. Zool., 1881, xxxv, p. fi32. 



^M^iii. publi6e8 Soc. philomat. Paris I'Occas. Cciitoii. Foiidatiou, 1888, p. 165. 



3Annal. de Microgr., 1890, ii, p. 196. ' 



■•Die Protozoen als Krankbeitserreger, 1891, 2 ed., pp. 19-24. 



fi Aniial. do Microgr.. 1892, iv, pp. 620-621. 



"Also Klciuellbe^g'^s li(|uid (Ileiincguy, 1888). 



' Hciinoguy (1888) also used ])icrocariiiin(!. • 



» Jouru. Anat. et Pbysiol., Paris, 1891, xxvii, pp. 398-100. 



