214 CHAPTER XVI. 



alcohol, containing at most O'l per cent, of hydrochloric acid, until hardly 

 any more colour comes away ; stain for one to three hours in gentian violet 

 ( 273) ; wash for a short time in distilled water ; treat with concentrated, 

 or at least fairly strong, aqueous solution of orange G, which " in virtue of 

 its acid properties " washes out most of the gentian. After at most a few 

 minutes, whilst pale violet clouds are still being given off from the sections 

 on agitation, bring them into absolute alcohol until hardly any more colour 

 comes away, clear in clove or bergamot oil, and mount in damar or balsam 

 before the last pale clouds of colour have ceased to come away. The orange 

 must be the orange G (last section). 



This is not a triple stain in the sense of giving three different colours in 

 the result ; it is a nuclear and plasrnatic stain in mixed tones ; the orange, 

 according to Flemming, does not act as a separate stain, but as an agent for 

 the differentiation of the gentian stain. I am unable to find that it acts in 

 this way, for I do not find that it washes out any of the gentian ; at all 

 events, in my preparations no perceptible clouds of colour come away (but 

 see last section). It seems to me more probable that it acts by forming a 

 " neutral " colour with the gentian (see next section and 262). 



See also FLEMMING in Arch. Anat. Phys., Anat. Abth., 1897, p. 175. 



Never popular, this clumsy and uncertain process is now little used. 



284. REINKE'S Modification of FLEMMING'S Orange Method (Arch, 

 f. mik. Anat., xliv, 2, 1894, p. 262). Sections of material fixed in liquid of 



HERMANN are put for twenty-four hours into a concentrated solution of 

 potassium sulphite. They are washed with water and stained for an hour 

 or two in safranin. They are then well washed with water, and stained for 

 twenty-four hours in a "neutral" mixture of gentian and orange, prepared 

 as follows : 



To a concentrated aqueous solution of gentian violet are added " a few 

 drops " of a like solution of orange G. The solution precipitates in part, 

 owing to the formation of an imperfectly soluble "neutral" colour; but 

 becomes almost clear again if an excess of water be added. A drop of the 

 mixture placed on blotting-paper should form a violet or brown spot with a 

 narrow orange border. The solution is not to be filtered, but the sections 

 are to be stained in it as it is (it is impossible to make out whether REINKE 

 means the undiluted mixture, or the mixture made almost clear by addition 

 of water). It is said that the "neutral" solution may be preserved for 

 future use by adding to it one third of alcohol. After staining, you differ- 

 entiate rapidly with alcohol, and clear with clove oil. 



I have tried this process, and obtained exactly the same results as with 

 Fleinming's process, and so have other workers. 



285. Metanil Yellow (Metanilgelb). See GHIESBACH (Zeit. f. iviss. 

 Mik., iv, 4, 1887, p. 448; see also Journ. Roy. Mic. Soc., 1889, p. 464). It 

 is said to have a certain affinity for various elements belonging to the group 

 of the connective tissues. 



286. Sauregelb (Echtgelb), Tropaeolin O., Crocein, Gold Orange 

 (see GrEIESBACH, Ari-fi.f. mik. Anat., xxii, p. 132). 



