Preparing a quality control material for blood gases, pH and hemoglobinometry using stroma-free hemoglobin solution

Abstract

Properties of a quality control material for blood gases and pH should be similar to normal human whole blood with respect to oxygen buffering and acid-base behaviour. A hemoglobin solution may potentially fulfill this. However, the drawbacks of such a solution are the high oxygen affinity (lowp₅₀), especially when it is prepared from human blood, and the improper concentration of bicarbonate.

Bicarbonate is added to human stroma-free hemoglobin solution (SFHS), prepared as described previously, to obtain the desired pH and pCO₂ combinations. Tonometry was used to determine the appropriate concentration of bicarbonate, which is 22.5 mmol/L to obtain an acidotic, and 29 mmol/L for both an alkalotic, and normal pH and pCO₂ combination. Inositolhexaphosphate (IHP) is added to SFHS containing bicarbonate to obtain a normal p₅₀ (around 3.55 kPa). Tonometry was used to determine the molar ratio of IHP/Hb₄ (mol/mol) at which this is achieved. The molar ratios of IHP/Hb₄ are 1.52, 1.74 and 3.40 for preparations with an acidotic, normal and alkalotic pH, respectively. In human SFHS n_Hill is 2.55 in the absence of IHP, n_Hill is at minimum 1.71 at a molar ratio IHP/Hb₄ of 1.86 and increases to 2.53 at a molar ratio IHP/Hb₄ of 5.04 and higher. Because the p₅₀ will decrease with xHi this was studied at molar ratios of IHP/Hb₄ of 0.2 and 4, which covers the range of ratios as used. At these molar ratios of 0.2 and 4, the decrease in p₅₀ is 0.017 kPa/%Hi, 0.023 kPa/%Hi and 0.028 kPa/%Hi, respectively.

Because bovine Hb has p₅₀ near that of normal human blood, it is also used. The oxygen affinity shows a small decrease (p₅₀ increases from 3.05 to 5.27 kPa) on addition of IHP. In the absence of IHP, n_Hill is 2.51 and n_Hill is at maximum 3.35 at molar ratios IHP/Hb₄ between 3.00 to 4.56. At higher molar ratios n_Hill decreases to 2.90.