and Laboratory Methods. 15(51 



by hardening in potassium bichromate, followed by impregnation with silver 

 nitrate. The production of the metallic deposit is not in the nature of a staining 

 process ; it is the outcome, rather, of an interaction between the potassium 

 bichromate and the silver nitrate employed, in conjunction with the substances 

 present in the nervous elements themselves. Since the chemistry of nervous 

 tissue is continually changing during life, it follows that the results of this method 

 will be far from uniform. In one animal, certain elements may be brought out 

 with all the crisp sharpness of a silhouette, while in another individual the 

 corresponding structures may not appear at all. The reactions of the tissues in 

 the two instances have differed sufficiently to thus affect the result. Hence the 

 apparent qapriciousness of the method is due to the presence of physiological 

 factors often quite unknown. The character of some of these physiological 

 conditions has been discussed by me elsewhere.* 



Freshness of tissue and active metabolism are factors indispensable to suc- 

 cess. A young animal will yield better results than an old one ; and one direct 

 from the field is preferable to one which has been kept for a time in confine- 

 ment. In any case, the parts of the nervous system to be studied should be 

 placed in the hardening mixture as soon as possible after the death of the 

 animal. 



a. The Rapid Method of Golgi. — Of the many schemes for securing the forma- 

 tion of the chrome-silver deposit, the one which has given me the largest returns 

 is the so-called " rapid " method of Golgi. For this purpose, the pieces should 

 be taken small, not over two or three millimeters in thickness for a slice of the 

 brain. Place these fresh slices directly in the hardening mixture : 



Potassium bichromate, 8.5 per cent. sol. - 4 vols. 



Osmic acid, 1 per cent. sol. . . . 1 vol. 



This reagent, while somewhat expensive, must be used liberally. There should 

 always be a large volume in proportion to the mass of the nervous matter present, 

 and the solution should be renewed before it shows the least signs of becoming 

 turbid. Hardening must take place in the dark. 



The duration of the hardening is the all-important factor which we have 

 under control. If the tissues be hardened for too short a time, the impregna- 

 tion will be diffuse ; while an overhardened specimen may permit of no impregna- 

 tion at all. The proper length of hardening must be determined by experiment 

 for each animal ; it will most often be found to lie between the limits of two to 

 three days where supporting elements are to be shown, three to five days for 

 neurones, and five to seven days for nerve-fibres. 



Impregnation is secured by bringing the hardened slices into a solution of 

 silver nitrate : 



Silver nitrate crystals . . . . 0.75 gram. 



Distilled water ----- 100.00 c. c. 



A solution which has been used once will answer for washing the specimens 



until the copious precipitate ceases to form. Then pour on a liberal quantity of 



the fresh solution, and change it frequently. It should not be allowed to take 



* Houser, 1901. The Neurones and Supporting Elements of the Brain of a Selachian: Jour- 

 nal of Comparative Neurology, Vol. XI. 



