300 VI. HEMOGLOBIN 



absorption band of pyridine hemochrome has been found rather variable 

 (619,629), and various authors have reported different intensity ratios of 

 the hemochrome absorption bands obtained from hemin and from hemoglobin 

 by pyridine (cf. Section 2.2.4.)- 



An instrument for clinical use has been designed by Holiday, Kerridge, 

 and Smith {1325), in which use is made of the Soret band of oxyhemoglobin. 

 The spectral region from 380 mju to 420 m^ is isolated by means of filters, 

 and measurement is made by means of a potassium photocell. The method 

 would include in the measurement hem/globin and sulfhemoglobin, but not 

 choleglobin. 



9.4. Colorimetric Methods 



While colorimetric methods are simple and cheap they are certainly 

 less exact than those previously described. On account of the com- 

 plicated absorption curves of most hemoglobin derivatives it is 

 impossible to devise artificial standards of exactly the same color. 

 One has therefore either to use solutions of hemoglobin deriv^atives 

 such as carboxyhemoglobin as standards, or, in order to use artificial 

 standards, the hemoglobin derivatives have to be transformed to 

 hematin derivatives of less selective absorption. In the first method 

 the stability of the standard is the weak point, and in the latter the 

 weakness lies in the need for chemical reactions which are not yet 

 perfectly understood {cf. Sections 2.4.2. and 2.4.3.) and which depend 

 on external conditions. 



The most commonly used method is the acid hematin method, in 

 which hemoglobin is transformed to "acid hematin" by hydrochloric 

 acid, and the brown color compared with that of a glass standard. 

 It has been criticized by several authors {cf. 158,2169) for more than 

 one reason. The solution is a colloidal one and the color development 

 depends on time and temperature, the plasma exerts some influence, 

 and the color varies with illumination. Nevertheless, although the 

 method is not exact, it gives quite satisfactory results for clinical 

 purposes, provided the conditions of color development and reading 

 are carefully controlled. 



Failure to maintain these conditions and the use of primitive and incor- 

 rectly standardized instruments and of the antiquated dilution method are 

 far more frequently to blame for wrong results than the method itself. There 

 is little excuse for this sloppy technique. As early as 1919 Newcomer {20Jf.8) 

 developed a satisfactory color disk for use in a colorimeter and defined the 

 conditions of color development. Recently a nomogram has been devised 

 (207) for correcting readings at various times after the addition of the acid 

 and at various temperatures to the value obtained after sixty minutes at 



