PRECISE DETERMINATION OF IRON ISOTOPE RATIOS IN WHOLE-BLOOD USING INDUCTIVELY COUPLED PLASMA MASS-SPECTROMETRY - INVITED LECTURE

The feasibility of measuring Fe absorption by incorporation of stable Fe isotopes into red blood cells has been assessed. The clinical protocol necessitated giving 10 mg of Fe-57 orally and 0.5 mg of Fe-58 intravenously, with only two blood samples needed, one basal and one after 14 d when most of the tracer is in the circulating red blood cells. Iron absorption was determined by comparison of Fe-57:Fe-56 and Fe-58:Fe-56 enrichments, which required great confidence in the reliability of Fe-56 measurements. Estimation of required precision by theoretical calculations suggested that a relative standard deviation of < 0.9% would be required to detect the enrichment in isotope ratios, which necessitated finding the optimum Fe signal in relation to the interfering polyatomic species present in both aqueous standards and whole blood samples diluted in buffer. It was found that Fe solutions of 10 ppm or greater routinely gave a precision of < 0.4% for Fe-57:Fe-56 and < 0.6% for Fe-58:Fe-56, approaching counting statistics. Assessment of dead time correction ensured that the concentration of Fe had a negligible effect on the isotope ratios, and bias correction, by running standards, ensured comparability within and across assay occasions, correcting for minor variations in blank subtraction for the less abundant isotopes. The use of a range of enriched aqueous and spiked whole blood samples showed that measured and calculated abundances correlated with a slope of unity. Blood samples from two subjects showed that after incorporation of enriched isotopes, isotope ratios of Fe-57:Fe-56 and Fe-58:Fe-56 were clearly distinguishable (standard deviation > 9) from the baseline. Inductively coupled plasma mass spectrometry with conventional aqueous sample introduction can be optimized to give precise measurement of all Fe isotope ratios in whole blood permitting clinical studies of Fe absorption.