1906.] Liver Cells to the Blood-vessels and Lymphatics. 457 



unnecessary in the case of the liver, and as a rale we found that we got 

 better injection preparations by using the carmine gelatine to drive out the 

 blood, but the cells of the liver can be injected quite well after a preliminary 

 injection of salt solution, and even after the injection of saline saturated with 

 chloroform. 



When the injection material was flowing freely from the opened inferior 

 vena cava the portal vein was ligatured. In some experiments we allowed 

 the injection material to fill up and distend the liver by ligaturing the 

 inferior vena cava above and below the diaphragm before tying the portal 

 vein, in others we left the inferior vena cava unligatured and allowed free 

 escape of the fluid all the time, so as to avoid any excess of pressure. In one 

 animal, a dog, the portal vein and hepatic artery were ligatured, and the 

 cannula was put into the aorta close to the heart, a pressure of 120 mm. Hg. 

 being used. There was a free escape all the time from the inferior vena cava 

 above the diaphragm, but, nevertheless, some of the injection mass passed 

 backwards along the hepatic veins and reached the central parts of the lobules 

 of the liver. Even under these circumstances the liver cells near the central 

 vein of each lobule were copiously injected with the carmine gelatine. The 

 pressures employed varied from 60 to 160 mm. of Hg when the injection was 

 made from the aorta, and rarely exceeded 20 mm. of Hg when made from 

 the portal vein. 



"We have also injected several frogs with carmine gelatine. In order to get 

 a free flow of the gelatine through the vessels, each frog was pithed and then 

 placed for half an hour in the warm bath (37° C), at the end of which time it 

 was sufficiently warmed to prevent solidification of the gelatine. 



In all cases, after the injection was completed, the liver was removed if the 

 animals were large ; if small, the abdomen and thorax were freely opened, and 

 the liver or the whole animal placed at once in 10-per-cent. solution of 

 formalin with some ice added. When the gelatine had set the liver was 

 removed, cut into pieces and put back into 10-per-cent. formalin. When 

 thoroughly fixed, pieces from different parts of the organ were dehydrated 

 with alcohol, cut in paraffin, and lightly stained with hematoxylin. If weak 

 haematoxylin be used the carmine retains its colour while the nuclei of the 

 liver cells give a good blue contrast. Deep staining must be avoided as it 

 stains the gelatine and masks the carmine. 



The use of two injection masses of different colour we found to be unsatis- 

 factory. We tried Prussian blue mass followed by carmine gelatine in one 

 dog, keeping up the first injection for some time so as to fill the lymphatics 

 of the liver. Carmine gelatine was then run through to wash away the 

 Prussian blue from the blood-vessels, and to fill them with red injection. 



2 M 2 



