34 FIXING AND HARDENING AGENTS. 



usual stains. So also EDINGER (Zeit. wiss. Mik., i, 1884, p. 126 ; 

 nitric acid 1 : 20 for five minutes). UNNA (Arch. Mik. Anat., xxx, 

 1887, p. 47) holds that the chrome is present in the tissues in the 

 form of chromic chromate, and removes it by treatment with per- 

 oxide of hydrogen. OVERTON (Zeit. wiss. Mik., vii, 1890, p. 9) 

 employs a weak solution of sulphurous acid, which converts it into 

 a sulphate. See also the directions for bleaching osmic acid prepara- 

 tions, 35. 



Tissues that have been fixed in chromic acid may be stained in 

 aqueous solutions, as water does not have an injurious effect on 

 them. 



The best stain for chromic material that has not been treated by 

 Mayer's special process, or by a similar one, is hsematoxylin, or, for 

 sections, the basic tar colours. 



Chromic acid is not a very penetrating reagent, and for this 

 reason, as well as for others, is now seldom used pure tor fixing. 



For prolonged hardening it is generally employed in strengths 

 of J to \ per cent., the immersion lasting a few days or a few 

 weeks, according to the size and nature of the object. Mucous 

 membrane, for instance, will harden satisfactorily in a few days ; 

 brain will require some six weeks. 



Large quantities of the solution must be taken (at least 200 grammes 

 for a piece of tissue of 1 centimetre cube Ranvier). 



In order to obtain the best results you should not employ portions 

 of tissue of more than an inch cube. For a human spinal cord you 

 should take 2 litres of solution, and change it for fresh after a few 

 days. Six weeks or two months are necessary to complete the 

 hardening. 



I think it is frequently useful to add a little glycerin ; there is less 

 brittleness. 



The solution should be taken weak at first, and the strength 

 increased after a time. The objects should be removed from the 

 solution as soon as they have acquired the desired consistency, as if 

 left too long they will become brittle. They may be preserved till 

 wanted in alcohol (95 per cent.). It is well to wash them out in 

 water for twenty-four or forty-eight hours before putting them into 

 the alcohol. After a time they generally become green in the 

 alcohol. They may be bleached if desired. 



Chromic acid is a most powerful and rapid hardening agent. 

 (By it you may obtain in a few days a degree of hardening that you 

 would hardly obtain in as many weeks with bichromate, for instance.) 

 It has the defect of a great tendency to cause brittleness. 



