a. Principle. Blood is diluted with a dilute solution of potassium ferricyanide and potassium cyanide at a slightly alkaline pH. The ferricyanide converts the hemoglobin to methemoglobin. The cyanide then reacts with the methemoglobin to form the stable cyanmethemoglobin. The color intensity is measured in a spectrophotometer at a wavelength of 540 mm. The optical density is proportional to the concentration of hemoglobin.
(1) Cyanmethemoglobin is the most stable of the various hemoglobin pigments showing no evidence of deterioration after 6 years of storage in a refrigerator. The availability of prepared standards is a distinct advantage of this technique. All hemoglobin derivatives are converted to cyanmethemoglobin with the exception of sulfhemoglobin.
(2) This method is highly accurate and is the most direct analysis available for total hemin or hemoglobin iron. Its disadvantage is the use of cyanide compounds, which, if handled carefully, should present little hazard.
(3) For accuracy in hemoglobin determinations, it is absolutely necessary that the spectrophotometer and Sahli pipets be accurately calibrated.
(4) Venous samples give more constant values than capillary samples.
(5) If the procedure is performed properly, the degree of accuracy is +2 to 3 percent.