ORIGIN OF BLOOD AND ENDOTHELIUM 11 



in picro-acetic (saturated aqueous solution of picric acid and 

 5 per cent glacial acetic) from thirty to forty minutes, then 

 put into 70 per cent alcohol. This is frequently changed in 

 order to wash out the picric acid. After they have been about 

 one-half hour in the 70 per cent alcohol, the egg membrane is 

 removed with fine dissecting needles. This is the most favor- 

 able time for removing the membrane. If the eggs have been 

 left for a long time in the alcohol, the membrane is more difficult 

 to remove and the embryo is brittle and more liable to injury. 



After removing the entire membrane, the yolk-sac is then 

 punctured at its ventral pole and the yolk mass very slowly 

 and cautiously removed from the sac. To remove the yolk 

 mass, requires a great deal of practice and extreme care in every 

 case. It should be done with the use of a binocular microscope 

 so that the operator can be certain not to tear or destroy the 

 yolk-sac or injure the delicate heart lying close above the yolk. 

 After a great deal of practice it is possible to remove the yolk 

 from a number of embryos and leave the yolk-sac in perfect con- 

 dition with the heart and pericardium practically undisturbed. 

 In the great majority of cases, however, it is generally impossible 

 to completely remove the yolk. It is usually necessary to remove 

 the yolk on account of the fact that when the eggs are imbedded 

 in either paraffin or celloidin, the yolk becomes so hard that it 

 often breaks the sections and makes it very difficult to get a 

 complete or perfect series. When the yolk-sac is punctured one- 

 half hour after having been in the 70 per cent alcohol, the yolk 

 material is in a gummy or viscid condition and is more easily 

 removed than at any other period tried. 



After having removed the egg membrane and the yolk, the 

 embryos are then allowed to stand twenty-four hours in 70 

 per cent alcohol when they are changed to 80 per cent to be 

 kept until the time of imbedding. The embryos are imbedded 

 in paraffin and cut in serial sections from five to ten micra thick. 

 They are stained in hematoxylin and eosin and extracted or 

 carefully differentiated so as to bring out a clear stain of the 

 blood cells and tissues. A complete series of these embryos have 

 been made from a time before the appearance of blood up to 



