Assay of creatinine using the peroxidase activity of copper-creatinine complexes

Objectives: It was our goal to develop a urine dipstick that could measure creatinine with a peroxidase reaction. The simultaneous measurement of albu min and creatinine permits the estimation of the 24-h albumin excretion, an important value in judging existing or likely development of renal failure . A highly sensitive dye-binding dipstick method for albumin exists, and a suitable dipstick for the assay for urine creatinine is described here. Methods: Copper-creatinine and iron-creatinine complexes have peroxidase ac tivity. With 3,3',5,5'-tetramethylbenzidine (TMB), and diisopropyl benzene dihydroperoxide (DBDH); the peroxidase activity of copper-creatinine and ir on-creatinine complexes can be demonstrated. This reaction was used in the assay of urine creatinine either in solution or by a suitably impregnated u rine dipstick. Results: Our method based on the peroxidase activity of the copper-creatini ne complex has an analytical range for creatinine of 100 mg/L (0.884 mmol/L ) to 3000 mg/L (26.52 mmol/L). The creatinine assay is free from most inter fering compounds that may be present in urine. Hemoglobin is an interferent , and its effects can be reduced but not eliminated by the addition of 4-hy droxy-2-methyl quinoline. We do not recommend using the dipsticks when visi ble blood is present or if the dipstick blood test is positive. The copper- creatinine complex oxidizes ascorbic acid; however, we were able to modify the reaction conditions so that ascorbic acid at < 4.4 g/L does not interfe re. We found good agreement on fresh urines between the creatinine dipstick results and those by a standard rate-Jaffe cuvet method for creatinine. Discussion: With the simultaneous measurement of creatinine and albumin in urine, the albumin/creatinine ratio can be determined effectively reducing or eliminating the occasional false-negative and false-positive result in t hose with dilute or concentrated urines, respectively. The dipstick test fo r these analytes permits the simple identification of individuals with poss ible albuminuria and could serve well in a point-of-care setting. Copyright (C) 2000 The Canadian Society of Clinical Chemists.