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CLIN. CHEM. 35/1, 134-138 (1989)
Effectsof Various Serum Proteinson Quantificationof Fructosamine Santiago Rodrlguez-Segade,1
Santiago Lojo, N. FelIx Caml#{241}a, J. Manuel Paz, and Rarn#{243}n Del RIo
We determined fructosamine concentrations with the CentrifiChem 600 centrifugal analyzer and the Hitachi 737 discrete analyzer. Reference intervals agreed with the most recently published results, and values in fasting patients were significantly correlated with glycated hemoglobin, plasma glucose, albumin, f3- and y-globulins, lgG, IgA, 1gM, and total protein. Partial correlation analysis showed that only fructosamine and IgA were dependently related. In a group of nondiabetic patients with pathological values for IgA concentrations, 79.4% had pathological values for fructosamine. These results throw doubt on the clinical value of fructosamine determinations if serum IgA is not taken into account.
Additional Keyphrases: reference interval variation,sourceof
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.
diabetes
IgA
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albumin
Glycated hemoglobin (more specifically, glycated HbA1) is an accepted measure of long-term average glycemia. Because nonenzymatic glycation also affects other proteins (1, 2), glycated albumin or total glycated protein (the “fructosamine assay”) can be used as measures of short-term (three-week) average glycemia (3-5). Of the various methodsfor determining fructosainine (3), the nitroblue tetrazolium colorimetric method of Johnson et al. (6) is the most
popular in clinical laboratories because of its speed, reproducibility, low cost, and ready automation (3, 5, 7-12). There is currently considerable controversy as to the relationship between the concentration of albumin or total protein and that of fructosamine. Some authors have reported that hypoalbuminemia detracts from the validity of the assay, but that measurements of fructosamine concentrations are independent of albumin concentrations when the latter exceed 30-35 gIL (4, 13-15). However, Van DieijenDepartment of Biochemistry and Serviceof Laboratory Central, Hospital General de Galicia, Faculty of Medicine, University of
Santiagode Compostela,Santiagode Compostela,Spain. ‘Address for correspondence: Department of Biochemistry, Faculty of Pharmacy,University of Santiagode Compostela,Santiago de Compostela,Spain. Received August 18, 1988; accepted October 11, 1988. 134
CLINICALCHEMISTRY, Vol. 35, No. 1, 1989
Visser et al. (16) found that fructosamine measurements are affected by the concentration of albumin, whether less or greater than 30 g/L, and suggested, as a correction, subtracting 0.023 mmol of fructosamine per gram of albumin per liter. The dependence of fructosamine concentrations on the concentrations of albumin and total protein has also been
reported by Mosca et al. (12) and by Howey et al. (9), who suggested using the correction: fructosamine (mmolIL) = 0.03 albumin (g/L) + 0.9 (mmol!L). Lim and Staley (14) did not find fructosamine measurements to be significantly affected by albumin concentration, but did point out that a reduction in protein half-life, caused by enhanced protein catabolism, would mean that the concentration of fructosamine would be reflecting mean glycemia over periods shorter than the expected 21 days. Lloyd and Marples (17), measuring fructosamine in thyrotoxic or hypothyroid patients, in whom protein turnover is respectively increased or reduced, found that fructosainine concentration was significantly decreased in the former and significantly increased in the latter. Though it is generally assumed that albumin makes the largest contribution to the total glycated plasma protein concentration, the relative importance of other plasma proteins in the fructosamine assay has never been investigated. Here we report evidence that immunoglobulin A (IgA) in plasma, especially at pathological concentrations, has a marked effect on fructosamine measurements.
Materials and Methods Analytical Procedures
Fructosamine assay. Serum fructosamine was determined by the method of Johnson et al. (6), as implemented with a CentrifiChem 600 centrifugal analyzer (Baker Instruments Ltd., Pleasantville, NY 10570) and a Hitachi 737 discrete analyzer (Hitachi Ltd., Tokyo, Japan) and the settings listed in Table 1. The mtroblue tetrazolium chloride (NBT) reagent was prepared in house, 250 zmol/L in 0.1 mol/L carbonate buffer (pH 10.35). To standardize results, we used the calibrator supplied in a fructosamine test kit (F. Hoffmann-La Roche, Basle, Switzerland).
Other assays. Glucose, albumin, and total protein concentration were determined in the Hitachi 737, with reagents supplied by Boehringer Mannheim GmbH, Mannheim, F.R.G. Proportions of Jib A1 were measured by isoelectric focusing on commercial polyacrylamide gels and quantified with an LKB 2202 Ultroscan laser densitometer equipped with an LKB 2220 recording integrator (LKB, Bromma, Sweden). IgG, IgA, and 1gM were determined by using an Array Protein System Analyzer and appropriate antisera (Beckman Instruments, Fullerton, CA 92621). Protein electrophoresis was performed on cellulose acetate strips with 90 mmol/L sodium barbital buffer (pH 8.6), and protein was stained with Ponceau S dye (Helena Laboratories, Beaumont, TX 77704). Subjects The reference
was established with the aid of 122 blood donors (64 men and 58 women, ages 17-72 y, median 37 y). The rest of the study was carried out with 197 outpatients: 66 diabetics undergoing various treatments and 131 patients with no history of diabetes and whose venous plasma glucose concentrations was
