The Colorimetric Determination of Hemoglobin.*

The method to be described for the determination of hemoglobin depends upon the comparison, in a calorimeter, of carbon monoxide hemoglobin solutions, one of which has a known hemoglobin content. Hoppe-Seyler (1) was the first to describe carbon monoxide hemoglobin and to make use of this stable combination for estimating the hemoglobin content of blood. He devised a “double pipette” for comparing the unknown carbon monoxide hemoglobin solution with the standard carbon monoxide hemoglobin solution, prepared from hemoglobin crystals; but the method never came into general use, because of the many technical difficulties involved. Haldane (2) suggested a much simpler method for comparing carbon monoxide solutions, using the apparatus employed by Gowers (3) for comparing oxyhemoglobin solutions with a picro-carmine standard. This apparatus was later employed by Sahli (4) who prepared an acid hematin standard by adding dilute hydrochloric acid to blood. A critical discussion of the various methods in use for the estimating of hemoglobin is beyond the scope of this paper. As Haldane (2) has pointed out, artificially colored solutions and tinted glass present great difficulties in standardization with a definite strength of hemoglobin solution. With a certain strength of color solution or tinted glass, it is possible to imitate quite perfectly the tint of a given hemoglobin solution provided the quality of light remains the same. Any variation from these standard conditions, either in quality of light or strength of hemoglobin in solution leads to serious errors. Haldane at-