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Mol Biotechnol. 2016 May;58(5):373-80. doi: 10.1007/s12033-016-9936-z.

Colorimetric Glucose Assay Based on Magnetic Particles Having Pseudo-peroxidase Activity and Immobilized Glucose Oxidase.

Author information

1
Faculty of Military Health Sciences, University of Defence, Trebesska 1575, Hradec Kralove, 500 01, Czech Republic.
2
Department of Chemical Drugs, Faculty of Pharmacy, University of Veterinary and Pharmaceutical Sciences, Brno, Czech Republic.
3
Laboratory of Structural Biology and Proteomics, Faculty of Pharmacy, University of Veterinary and Pharmaceutical Sciences, Palackeho tr 1/1946, 613 00, Brno, Czech Republic. kruzliakpeter@gmail.com.
4
Institute of Geotechnics, Slovak Academy of Sciences, Kosice, Slovak Republic.
5
Faculty of Military Health Sciences, University of Defence, Trebesska 1575, Hradec Kralove, 500 01, Czech Republic. miroslav.pohanka@gmail.com.

Abstract

Magnetic particles (MPs) are currently used as a suitable alternative for peroxidase in the construction of novel biosensors, analytic and diagnostic methods. Their better chemical and thermal stabilities predestine them as appropriate pseudo-enzymatic catalysts. In this point of view, our research was focused on preparation of simply and fast method for immobilization of glucose oxidase onto surface of MPs with peroxidase-like activity. Spectrophotometric method (wavelength 450 nm) optimized for glucose determination using modified MPs has been successfully developed. Concentration curve for optimization of method was assayed, and Michaelis-Menten constant (K m) calculated, maximum reaction rate (V max), limit of detection, and correlation coefficient were determined to be 0.13 mmol/l (2.34 mg/dl), 1.79 pkat, 3.74 ┬Ámol/l (0.067 mg/dl), and 0.996, respectively. Interferences of other sugars such as sucrose, sorbitol, deoxyribose, maltose, and fructose were determined as well as effect of substances presenting in plasma (ascorbic acid, reduced glutathione, trolox, and urea). Results in comparison with positive and negative controls showed no interferences of the other sugars and no influence of plasma substances to measuring of glucose. The constructed method showed corresponding results with linear dependence and a correlation coefficient of 0.997. Possibility of repeated use of modified MPs was successfully proved.

KEYWORDS:

Glucose oxidase; Glucose sensor; Magnetic particles; Mimic peroxidase; Spectrophotometry

PMID:
27041274
DOI:
10.1007/s12033-016-9936-z
[Indexed for MEDLINE]

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