HEMOGLOBIN DETERMINATION 209 



of acid, in order to give a dilution having the most satisfactory 

 color for comparison with the standard. After the sample is added 

 to the acid, the mixture must be allowed to stand, preferably in 

 a warm place, for at least ten minutes for the full color to develop. 

 Readings taken sooner will be too low. 



In cold weather, hemolysis and color development take place 

 more slowly, and the application of gentle heat will hasten the 

 process. As a routine procedure, immersion of the tube in a warm 

 water bath is recommended. The color comparison may be made 

 in either the Autenrieth-Hellige or the Duboscq colorimeter with 

 a standard acid hematin solution. The average of at least several 

 readings is taken. The calculation is simple and described below 

 for each instrument. 



Standard and Calculation for the Duboscq Colorimeter. The 

 standard for comparison is a 0.5 per cent blood solution, which is 

 set at 10 upon the Duboscq scale. Hence the per cent hemoglobin = 



1.5X10X100 T , . , . ,. . r , ,. , 



r . It is desirable to make an actual calibration of 



Reading 



the instrument for the solutions to be examined. 



Standard and Calculation for the Autenrieth-Hellige Colorim- 

 eter. The standard for comparison is a 1.0 per cent blood 

 solution. Experience has pointed to the necessity for different 

 concentrations of standard solutions in each kind of instrument 

 in order to secure most satisfactory color comparisons. It may 

 be found that the scale on the Autenrieth colorimeter may be 

 inaccurately placed. There is another source of error that cannot 

 be corrected by a mere resetting of the scale. The glass wedge 

 containing the standard is not mathematically perfect; therefore 

 for accurate work, this wedge containing the 1 per cent standard 

 should be calibrated against solutions of known amounts of blood 

 in 0.1 N HC1 contained in the small cup. Thus a curve may be 

 constructed from which may be read at once the percentage 

 hemoglobin corresponding to a given reading in the scale. This 

 calibration takes a short time, and holds good for that wedge and 

 instrument as long as other conditions are maintained. 



For the conditions given here, if the instrument were perfectly 



, , , ,, , , ,. ,, , 3.0X10X100 



constructed, the calculation would be =r- r . =per cent 



Reading 



hemoglobin, but this relation holds for only a portion of the scale 

 (between 3.0 and 8.5). 



