STUDIES IN THE REGENERATION OF BLOOD 241 



mental use and during all of the time when they were not under direct 

 observation. This eliminated complications arising from having the 

 animals eat such complex mixtures as they would, had they been fed 

 table scraps. 



Technique. The preliminary experiments were designed to discover 

 the limit of successful bleedings in the rat, the period for regeneration 

 of blood after hemorrhages, and to find the effect of the dietary factors 

 on such regeneration. 



The rats on which hemorrhages were produced were kept in iron-wire 

 cages with paraffined tin cups for water and porcelain cups for food. 

 The methods and technique employed have been described by Miss 

 Ferry (1). The standard synthetic diet developed by the work of 

 Osborne and Mendel (2), (3), (4) and others was used as a basis for 

 all the dietaries employed. Adult and young animals of both sexes 

 were used, varying in average weight from 100 grams for the young 

 to 300 grams for the adults. The tables of Donaldson (5) were carefully 

 followed in estimating the blood volume of each experimental animal 

 used. 



Experimental technique. The blood of every rat was carefully ex- 

 amined for the number of erythrocytes and percentage of hemoglobin 

 before any hemorrhages were made. The counts were checked in every 

 case during the first 2 months until the operator had attained sufficient 

 skill to make such duplications unnecessary. The method of blood 

 count was the familiar one with a Thoma-Zeiss hemocytometer and the 

 use of either Hayem's or Toisson's solution for blood dilution. Hemo- 

 globin was determined by the colorimetric procedure of Cohen and 

 Smith (6) . Rat blood was used as a standard and its color was checked 

 by fresh material at frequent intervals. 



The sample of blood for examination was taken from a drop extruded 

 when the end of the tail was snipped. The rat was placed in an open 

 glass bottle, the bottom of which had been removed and a tin cap with 

 a hole for the tail placed over the opening; leather straps from the neck 

 of the bottle held the rat securely in this armor (see fig. 1). This 

 arrangement allowed free manipulation of the tail by the experimenter. 

 The skin of the tail was softened with mineral oil. Previous to removing 

 the end of the rat's tail it was placed in warm water or brushed over 

 with toluene in order to suffuse the blood vessels. There are four blood 

 vessels in the rat's tail, one dorsal, one ventral and two lateral. The 

 blood was obtained from these by manipulating the tail after the fashion 

 of milking and allowing the fluid thus "milked" out to drop into a 



