Comparison of Bathophenanthroline Sulfonate and Ferene as Chromogens in Colorimetric Measurement of Low Hepatic Iron Concentration

Abstract

Sensitive and accurate measurement of hepatic iron concentration (HIC) is required to investigate liver fibrogenesis (1) and its influence on the outcome of interferon therapy for chronic viral hepatitis C (2)(3). Hepatic iron content can be measured by a quantitative chemical method and/or evaluated by semiquantitative histologic scoring. Quantitative chemical methods assess all liver iron forms, whereas histologic scoring evaluates only the hemosiderin form. A colorimetric method using bathophenanthroline sulfonate as chromogen was recommended in 1978 by the International Committee for Standardization in Hematology (ICSH) for determination of serum iron (4)(5). It was adapted by Barry and Sherlock (6) to the determination of HIC, and we recently evaluated it for measurement of low HIC (7). In 1990, the ICSH replaced bathophenanthroline sulfonate with ferene, a more sensitive chromogen, in the determination of serum iron (8). The aim of the present study was to evaluate the replacement of bathophenanthroline sulfonate with ferene to improve the sensitivity of the colorimetric determination of low HIC. We used samples of a frozen Wistar rat liver for quality control and determination of reliability criteria of both assays. We compared the results obtained with the two chromogens on 66 liver biopsies from patients with chronic liver diseases hospitalized in the Department of Hepatogastroenterology of the Pitie-Salpetriere Hospital. The clinical diagnoses of these patients are summarized in Table 1⇓ . We determined the CV for HIC measurements on two separate samples from the same liver specimen for each chromogen on 38 human liver biopsies. Histologic iron scoring was according to the method of Deugnier et al. (9); among the 66 biopsies, 20 had no stainable iron (score of 0), and the 46 others exhibited iron overload (score ≥6). Biopsies were fixed in 40 g/L formaldehyde and embedded in paraffin as part of …