New enzymatic assay of iron in serum.
A new enzymatic method for assaying iron in serum samples, suitable for automated analyzers, is reported. Three reagent mixtures are used: dilution buffer (pH 3.0; ascorbate), reagent 1 (pH 6.7; apoaconitase), and reagent 2 (pH 7.7; citrate, magnesium, and isocitrate dehydrogenase). Sera are diluted with dilution buffer. Fe3+ is liberated from transferrin in sera under acidic conditions, and then reduced by ascorbate. Reagent 1 is added to diluted specimens, and apoaconitase is reactivated by Fe2+ at neutral pH. The resulting solutions are mixed with reagent 2, so that holoaconitase hydrolyzes citrate to isocitrate and the isocitrate and NADP+ are converted to 2-oxoglutarate, NADPH, and CO2. Serum iron is determined linearly up to 70 mumol/L, with within-run CVs < or = 2.4% and day-to-day CVs < or = 2.9%. This method (y) gives results correlating with those of a Reference Method (x) proposed by the International Committee for Standardization in Haematology: y = 0.98x + 0.38 mumol/L (n = 72, r = 0.996, Sylx = 0.63 mumol/L). The mean (+/- SD) serum iron concentrations measured by our method were 18.5 +/- 5.4 and 15.2 +/- 6.0 mumol/L for 63 males and 166 females, respectively.
Fujita
T
T
Biochemical Research and Development Center, Oriental Yeast Co., Ltd., Osaka, Japan.
Hamasaki
H
H
Furukata
C
C
Nonobe
M
M
D000154:Aconitate Hydratase / Q000378:metabolism*; D001051:Apoenzymes / Q000378:metabolism; D001322:Autoanalysis; D002951:Citrates / Q000494:pharmacology; D019343:Citric Acid; D005260:Female; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007202:Indicators and Reagents; D007501:Iron / Q000097:blood*; D007521:Isocitrate Dehydrogenase / Q000378:metabolism*; D007700:Kinetics; D008274:Magnesium; D008297:Male; D012016:Reference Values; D012680:Sensitivity and Specificity; D014168:Transferrin / Q000378:metabolism
D001051:Apoenzymes; D002951:Citrates; D007202:Indicators and Reagents; D014168:Transferrin; D019343:Citric Acid; D007501:Iron; D007521:Isocitrate Dehydrogenase; D000154:Aconitate Hydratase; D008274:Magnesium