INTRODUCTION TO VITAL COLOURING 279 



to be flocculated by salts (p. 290). It is usual to give several in- 

 jections at intervals of hours or days and subsequently to remove 

 fragments for study while still alive in a saline medium. 



Another method is to mix the dry dye with the food. The fact 

 that bones become coloured if an animal eats madder appears to 

 have been known to the Chinese of antiquity/^ but this should 

 be regarded as an example of pseudovital colouring. Lipid- 

 soluble colouring agents may be dissolved in edible fats. The 

 method is simple but has the disadvantage that dosage cannot be 

 easily controlled. 



Yet another method is to remove loose cells or minute fragments 

 of tissue from the body and place them in a solution of the dye in 

 a saline fluid of the right osmotic pressure. A concentration of 

 0-01% or less is usually best. It is convenient to keep stock solu- 

 tions at 1% or h% in distilled water and to dilute these with the 

 appropriate saline solution just before use. Michaelis ^^^ called 

 this important method of vital colouring Die postmortale Fdrhiing, 

 but there is, of course, no reason why the animal from which the 

 cells were taken should die. Arnold ^^' ^^ made a careful study of 

 the dyeing of uherlehender cells. He chose this very suitable term 

 because the cells, having been removed from the body, survived 

 during the period of coloration. This should properly be called 

 survival or supervital dyeing (see p. 329). 



If one prefers to leave cells in their natural medium during the 

 process of supervital dyeing, it is possible to dissolve the dye 

 directly in the natural medium and thus avoid the use of any 

 artificial saline solution. To do this, the dye is dissolved in absolute 

 ethanol, and the solution spread on microscopical glass slides and 

 evaporated to dryness. ^^^ When a drop or two of blood or any 

 other natural fluid that contains cells is placed on a slide that has 

 been treated in this way, the dye dissolves in the fluid and colours 

 the living cells. It is desirable to take precautions to ensure that the 

 ethanol is not contaminated by anything that could damage the cells. 



A routine microscopical preparation of dead tissues focuses the 

 observer's attention on the distribution of chromatin. The appear- 

 ance of cells after vital dyeing is entirely diflPerent. Usually the 

 nucleus and ground cytoplasm are scarcely or not at all affected, 

 and attention is focused on certain cytoplasmic inclusions, the 

 existence of which is not even suggested in the routine slide. The 

 principal inclusions that are coloured are certain vacuoles, lipid 

 globules, and mitochondria. The vacuoles that are coloured are of 



