Iron and Total Iron-BindingCapacity in Serum of ... - Clinical Chemistry

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only in patients who had received an Intravenous Fe-dextran. (lmferon#{176})injection two to three days beforeblood sampling but could be generated In vitro by ...

CLIN. CHEM.

36/10, 1812-1815 (1990)

Iron and Total Iron-Binding Capacity in Serum of Patients Receiving Iron-Dextran: Kodak Ektachem Methodologies, Spectrophotometry, and Atomic-AbsorptionSpectrometry Compared N. Vercammen,

W.

Go.dhuys, A. Boeyckens,

R. Do

Roy,J. Sennesael,1 C. Sevens, and F. Gorus2

We compared

the analytical performance of the Kodak Ektachem XR700 assays of iron (Fe) and total Iron-binding capacity (TIBC) with that of a conventional ferrozine assay (performed with a Cobas-Blo) and occasionally with that of atomic-absorption spectrometry (AAS). The correlation was modest between Kodak and Cobas-Bio concentrations of Fe in serum (r = 0.79). Multiple outliers were noticed in samples from hemodialysis patients, with Cobas values exceeding those of Kodak by as much as 26 tmol/L. These intermethod differences were not dissipated by dialysis, but were invariably accompanied by an even higher total Fe content of the serum as judged by AAS (20 to 80 ,umoVL higher). TIBC values by Kodak and Cobas-Blo were highly correlated (r = 0.99); agaIn, the Cobas-Blo results exceeded those by Kodak In some hemodialysis patients, but always for samples with higher Fe concentrations by the Cobas-Bio. By AAS, the TIBC values of these patients also exceeded those by Kodak, to about the same extent as observed for serum Fe. These intermethod differences in Fe and TIBC were seen only in patients who had received an Intravenous Fe-dextran (lmferon#{176}) injection two to three days before blood sampling but could be generated In vitro by adding Imferon to serum from normal controls. Less than 6% of dextran-bound Fe is measured as Fe by Kodak, as opposed to 20-30% by Cobas-Bio and 89-120% by AAS. We conclude that the Kodak Fe slides are superior to liquid reagents, by exclusively measuring protein-bound circulating Fe pools. AddItional Keyphrases: parenteral iron Therapy

hemodiatysis

.

an#{216}ytic&error

In the past decade, the Ektachem multilayer film technology (Eastman Kodak, Rochester, NY) has found widespread application in clinical laboratories (1,2). The compartment.alization of dried reagents in juxtaposed thin, solid layers leads to generally longer stability, excellent reproducibility, and reduced susceptibility to interferences (1-6). Recently, Ektachem slides for assaying iron (Fe) and total iron-binding capacity (TIBC) in serum became commercially available (6). Here we evaluate their analytical performance in an Ektachem 700XR analyzer (Eastman Kodak) in comparison with conventional liquid reagents (7) adapted to a Cobas-Bio instrument (Hofl’mnnn-LaRoche, Basel, Switzerland) and, in selected conditions, to atomicabsorption spectrometry (AAS) (8).

Departments of Clinical Chemistry and’ Nephrology, Akademiach Ziekenhuis, Vrije Universiteit Brussel (AZ-VUB), Laarbeeklaan 101, B-1090 Brussels, Belgium. 2Address correspondence to this author at AZ-VUB. 3Nonstandard abbreviations: TIBC, total iron-binding capacity UIBC, unsaturated iron-binding capacity; and AAS, atomic absorption spectrometry. Received March 8, 1990; accepted July 5, 1990. 1812

CLINICALCHEMISTRY, Vol.36, No. 10, 1990

MaterIals and Methods Blood Samples We investigated both hospitalized subjects and ambula tory patients for whom an Fe and TIBC determination wa ordered. Blood was collected in dry plastic tubes (Sarstedt Haasrode, Belgium), and in some patients, in a lithiun heparin-coated tube (Sarstedt) as well. Samples were cen trifuged within 2 h after venipuncture, then divided intc aliquots, and all but one were stored at -20 #{176}C for furthei study. The remaining aliquot was kept at room tempera tore until assayed for Fe and TLBC later on the same day In selected uremic patients, blood was sampled before anc after hemodialysis or administration of parenteral Fedextran (Imferon#{174}; Fisons, Loughborough, U.K.). Iron Assays Ektachem Fe slides were used with an Ektachem XR7O( analyzer. Briefly, Fe3 is freed from transferrin at pH 4 anc subsequently reduced to Fe2 by means of p-meth. ylaminophenol sulfate. Finally, Fe2 reacts with N. {4 - [2,4 - bis(1,1 - dimethylpropyl)phenoxy]butyl - 5- meth. oxy - 6[2,3,6,7 tetrahydro-IH,5H,5H,8H - benzo()quinoli. zin-9yl]azo)-3-pyridine sulfonamide, a pyridyl-azo dye, tc form a red complex that is quantified by reflectance spec. trophotometry at 600 nm (6). We compared the Ektachem methodology with a fer. rozine method, using commercial liquid reagents (J. T Baker, Deventer, The Netherlands) adapted to a Cobas-Bic centrifugal analyzer (Hoffmann-LaRoche) according to th#{128} manufacturer’s specifications (7). The liquid reagents dil fered from the “dry chemistry” slides in that ferrozine wes used as chromogen and ascorbic acid (85 mmol/L) as reduc tant and the absorbance of the Fe2-ferrozine complex w monitored at 562 nm by conventional spectrophotometry. In selected sera, Fe concentrations were also determin by AAS (8) with an IL 457 instrument (Instrumentatio Laboratory, Analytical Instrument Division, Wilmington MA). The instrument was calibrated against Fe stan (9.1-45.4 MmolJL) prepared by diluting a commercial call brator (Atomic Spectral Standard, 1000 ppm, cat. no. 692 1; J. T. Baker) with a physiological salt solution (Stan 14ONaJ5K, cat. no. 35140; Instrumentation Laboratory For assays of Seronorm Trace Elements quality-contro serum (Nycomed, Oslo, Norway), the day-to-day CV w 3.4% (n = 6) at a mean Fe concentration of 22.6 mo (target value 23.0 mol/L). Analytical recovery of iro standards added to sera was 100-106%. The Kodak and Cobas-Bio methods were calibrated wi Kodak Calibrator/Diluent Set and Sigma Iron Stan (cat. no. 565-5; Sigma Diagnostics, St. Louis, MO), tively. Assays of commercial quality-control sera (Koda from Kodak and Seronorm from Nycomed, respectivel showed

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