Authors:
Jan Přibyl
and
Petr Skládal
Affiliation:
National Center for Biomolecular Research, Masaryk University, Czech Republic
Keyword(s):
Glycated hemoglobin, aminophenylboronic acid, biosensors, quartz crystal microbalance, heterogeneous affinity assay, microtitre plate.
Related
Ontology
Subjects/Areas/Topics:
Biomedical Engineering
;
Biomedical Instruments and Devices
;
Biomedical Sensors
;
Devices
;
Health Monitoring Devices
;
Human-Computer Interaction
;
Physiological Computing Systems
Abstract:
Development of bioanalytical assays for determination of glycated hemoglobin content in blood samples is reported. First, a combined biosensor setup for determination of total and glycated hemoglobin content was successfully developed and tested. The effect of various operating parameters, such as ionic strength, flow rate and instrumental set-up was optimized. The total hemoglobin content was analyzed by measuring of absorbance of the hemoglobin-cyanide derivative at 540 nm. Only one standard (calibrator), diluted in various proportions, was necessary for the method calibration. The full range of HbA1c content (4 to 15 %) presented in blood can be analyzed. Only 1 μl of blood was required for analysis. The developed method was successfully evaluated for analysis of blood samples collected from diabetic patients. Next, the heterogeneous affinity assay performed in a microtitre plate with an immobilized boronic acid is described. This assay is based on ELISA (Enzyme-Linked Immunosorbe
nt Assay) principle; however stable chemiselective ligand is used in this case. The content of glycated hemoglobin is determined according to its peroxidase activity after attachment to immobilized boronic acid derivative; the total hemoglobin concentration is measured as an absorbance at 405 nm.
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