THE MYXOSPORIDIA, OR PSOROSPERMS OF FISHES. 1D i 
VII.—MICROSCOPIC TECHNIQUE. 
The older observers used no reagents beyond acetie acid, potassium 
hydrate, etc. Biitschli! was the first to use a staining reagent. He 
believed that alum carmine stained nuclei in the ectoplasm. The 
first observer to employ modern technique was Henneguy.? Subse- 
quently Thélohan* employed similar technique, and Pfeiffer* devotes 
some space to the technique of protozoan investigation. Finally Hen- 
neguy and Thélohan® 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, paraffining, 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 the different stages of development. 
Culture in the blood (overhanging drop method) is recommended by 
Pfeiffer 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. Satranin, by Henneguy’s 
method,* evinces an electivity valuable in the study of development 
where we have to do with the most complex phenomena of cellular life 
under circumstances in which the small size of the elements renders 
observation extremely difficult. 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 effected by sulphuric acid (cold, 
concentrated). 
Vacuole: Best shown by very dilute iodine water (with potassium 
iodide). 
1 Ztschr. f. wiss. Zool., 1881, XXXvV, p. 632. 
?>Mém. publiées Soc. philomat. Paris l’Occas. Centen. Fondation, 1888, p. 165. 
3 Annal. de Microgr., 1890, 11, p. 196. 
‘Die Protozoen als Krankheitserreger, 1891, 2 ed., pp. 19-24. 
5 Annal. de Microgr., 1892, Iv, pp. 620-621. 
6 Also Kleinenberg’s liquid (Henneguy, 1888). 
7Henneguy (1888) also used picrocarmine. 
8 Journ. Anat. et Physiol., Paris, 1891, xxvil, pp. 398-400. 
