METALLIC STAINS 207 



allowed to cool (Ranvier's Traite, p. 826). They remain in 

 this until thoroughly impregnated (muscle twenty minutes, 

 epidermis two to four hours) ; reduction is affected either by 

 daylight in acidulated water, or in the dark in dilute formic acid 

 (1 part of the acid to 4 parts of water). 



405. Ranvier's Lemon-juice Method {Traite, p. 813). Ranvier 

 finds that of all acids lemon juice is the least hurtful to nerve- 

 endings. He therefore soaks pieces of tissue in fresh lemon 

 juice until they become transparent (five or ten minutes in the 

 case of muscle). They are then rapidly washed in water, brought 

 for about twenty minutes into 1 per cent, gold chloride solution, 

 washed again in water, and brought into a mixture of 50 c.c. of 

 distilled water and 2 drops of acetic acid. They are exposed to 

 the light for twenty-four to forty-eight hours. The preparations 

 thus obtained are good for immediate study, but are not per- 

 manent, the reduction of the gold being incomplete. In order to 

 obtain perfectly reduced, and therefore permanent, preparations, 

 the reduction should be done in the dark in a few cubic centi- 

 metres of dilute formic acid (1 part acid to 4 of water), which takes 

 about twenty-four hours. 



406. Viallane's Osmic Acid Method {Hist, et Dev. des Inseects, 

 1883, p. 42). The tissues are treated with osmic acid (1 per 

 cent, solution) until they begin to turn brown, then with 25 per 

 cent, formic acid for ten minutes ; they are then put into solution 

 of chloride of gold of 1 : 5000 (or even much weaker) for twenty- 

 four hours in the dark, then reduced in the light in 25 per cent, 

 formic acid. Lee found this an excellent method. 



Kerschner {Arch. inik. Anat., Ixxi, 1908, p. 522) puts till 

 brown into a mixture of 10 parts 5 per cent, formic acid with 1 

 part 2 per cent, .osmic acid, washes, puts for two to six hours 

 into 1 per cent, gold chloride in the dark, washes, puts for twelve 

 hours into 25 per cent, formic acid in the dark and then for 

 twenty-four in the light, and mounts in 50 per cent, glycerin 

 with 1 per cent, of formol. 



407. Other Methods. The numerous other methods that have 

 been proposed differ from the foregoing partly in respect of the 

 solutions used for impregnation, but chielly in respect of details 

 imagined for the purpose oi facilitating the reduction of the gold. 



Thus Bastiax employed a solution of gold chloride of a strength 

 of 1 to 2000, acidulated with HCl (1 drop to 75 c.c), and reduced 

 in a mixture of equal parts of formic acid and water kept warm. 



Hexocque {Arch, de VAnat. et de la Physiol., 1870, p. Ill) 

 impregnates in a 0-5 per cent, solution of gold chloride, washes 

 in water for twelve to twenty-four hours, and reduces in a nearly 

 saturated solution of tartaric acid at a temperature of 40° to 50° C. 

 Reduction is effected very rapidly, sometimes in a quarter of an 

 hour. 



