182 DETEKMINATION OF HAEMOGLOBIN. [BOOK I. 



Determination of Haemoglobin in the Blood. 



i. Hoppe- This method consists in comparing the tint of the 



blood diluted with a known volume of distilled water, 



with the tint of a solution of pure haemoglobin of 



known strength, and then adding water to the first until it assumes 



exactly the colour of the second. 



This method necessitates in the first place a solution of pure Oxy- 

 haemoglobin. Oxy-haemoglobin, prepared from the blood of the dog 

 and at least twice crystallized, is dissolved in water at C., and the 

 saturated solution is filtered. 50 c.c. are measured out in a capsule 

 and evaporated to dryness, first over a water-bath and then over 

 sulphuric acid, in vacua. In this way the strength of the solution is 

 determined. 



In addition to this solution there are needed two haematin- 

 .ometers 1 (see Fig. 16, p. .92), and an accurate burette divided into 

 tenths of a cubic centimetre. 



The two haematinometers being placed side by side, with a sheet 

 of white paper beneath them, and in such a position as to be 

 illuminated in exactly the same manner, 10 c.c. of the standard 

 solution, diluted with from 10 to 60 c.c. of water, are placed in the one 

 haematinometer. 



A solution, of known strength, of the blood to be investigated is 

 now made, e.g. by diluting 5 grammes of blood to 100 c.c.; 10 c.c. 

 of the solution are placed in the second haematinometer. 



The solution of blood will now be seen to be very much deeper 

 in tint than the solution of haemoglobin. Water is added to the 

 former, from a burette, until there is no perceptible difference in tint 

 between it and the standard solution. When this result has been 

 obtained, the amount of haemoglobin in the two solutions must 

 be equal. 



The method and the calculation required will be understood from the 

 following experiment quoted from Hoppe-Seyler : 20 '186 gnus, of denbrin- 

 ated blood were diluted with water to a volume of 400 c.c. 10 c.c. of 

 this solution were placed in a haematinometer and 38 c.c. of water had to 

 be added so as to produce a solution of the same shade as the standard 

 solution of haemoglobin which had been placed in a second haematinometer. 

 The volume of water which the whole quantity of the solution of blood 

 would have required to bring it to the standard tint is found by the 

 proportion 



10 : 38 :: 400 : x 

 # = 1520 c.c. 



1 Haematinometers similar to the one represented in Fig. 16 are constructed by 

 the opticians Schmidt u. Haensch, Stallschreiber-Strasse 4, Berlin ; they are sold in 

 pairs, at 30 marks (1. 10s.) the pair. 



