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KMID : 0385520100230060537
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Analytical Science & Technology
2010 Volume.23 No. 6 p.537 ~ p.543
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Bioconjugation by dual heterobifunctional coupling method : Use of the conjugates for the detection of dopamine
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Ryu Ji-Eun
Lee In-Sook
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Abstract
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Dopamine (DA) is an important neurotransmitter molecule of catecholamines. Its deficiency could lead to brain disorder such as Parkinson"s disease and schizophrenia. Therefore, it is necessary to establish a suitable analytical technique with sensitivity and simplicity. A competitive enzyme-linked immunosorbent assay for DA has been optimized and characterized. Assay sensitivity is controlled by two factors in competitive immunoassay. One is a nature and concentration of competitor, and the other is those of binder, antibody. Thus, optimization was performed: BSA-DA conjugate and antibody-avidin conjugate were prepared by dual heterobifunctional coupling method using SATA and SMCC. Assay condition was optimized with 6.66 ㎍mL?1 of BSA-DA and 4.17×10?10 M of antibody-avidin conjugate. A dose-response curve was constructed, and a limit of detection and a dynamic range for DA were accomplished to 2.3×10?2 ㎍ mL?1 and four orders of magnitude (1.0×10?7 M to 1.0×10?3 M), respectively. Calibration curve was constructed on dynamic range and least-squares regression of this data gave the following relationship: absorbance = -0.1098 log[DA]+0.0353 (R² = 0.9956).
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KEYWORD
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immunoassay, dopamine, heterobifunctional coupling, avidin/biotin
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