Journal of Applied Microscopy. 



541 



Histological Fixation by Injection. 



The necessity for fixing and preserving a large range of material for the courses 

 in Comparative Histology in Leiand Stanford Jr. University has led to the develop- 

 ment of a simple method of in toto fixation, which has thus far most admirably 

 answered every demand made upon it. For several years I have used this 

 method upon all sorts of vertebrate material, such as Myxinoids (Bdellostoma), 

 Selachians, Teleosts, Reptiles, Birds, and Mammals, and uniformly it has given 

 the best of results. The ideal histological fixation would be to surround each 

 individual cell with the killing liquid, and this can be practically obtained by 

 injection and by injection only. The apparatus used 

 must not be attacked by the fixing agents, and pre- 

 cautions must be taken to avoid such vascular con- 

 traction as would preclude the free access of the liquid 

 to every part. 



The apparatus in use here is extremely simple, con- 

 sisting essentially of two glass funnels, two pieces of 

 rubber tubing six or eight feet long, and two or three 

 of shorter lengths, a Y glass tube, four pinch- cocks, 

 and a supply of glass cannulas of various sizes. 



The accompanying cut shows the arrangement of 

 these parts. The long rubber tubes A, B, bearing 

 funnels at their free ends, are joined together by the Y 

 tube E, a simple pinch-cock being placed upon each im- 

 mediately above f. The third arm of the Y tube is 

 connected with a short rubber tube H, bearing the 

 cannula /. Another pinch-cock, sometimes convenient, 

 may be placed upon H. To facilitate the removal of 

 bubbles of air a T tube, K, bearing a short rubber tube, 

 C , with pinch-cock, should be placed near the lower end 

 of the tube a. 



The method of manipulation is extremely simple, 

 varying but slightly for different animals. If, for 

 example, a kitten is to be injected, the tube B is filled 

 through its funnel with normal saline solution warmed 

 to body temperature. The other funnel and tube A 

 are filled with the fixing agent, e. g., Zenker's fluid, 



warmed to the same degree. To the normal salt solution is added a few drops of 

 lactic acid, amyl nitrite, or some similar reagent to insure dilation of the blood vessels 

 and capillaries. The animal having been anaestheticised, the thorax is rapidly 

 opened, the apex of the heart is cut across, and the cannula is introduced through 

 the left ventricle into the ascending aorta and firmly tied. Before introducing the 

 cannula the salt solution is allowed to run through the whole apparatus below the 

 pinch-cock F, in order to avoid all bubbles of air. The pinch-cock on H is now 

 opened and the salt solution is injected through the circulatory system, washing 



