Bioluminescence immunoassay for neurotransmitter, serotonin using aequorin as a Label
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A sensitive competitive heterogeneous bioluminescence immunoassay for serotonin was developed using photoprotein, aequorin as a label for the first time with the optimal assay conditions; especially, serotoninavidin conjugate was prepared by Mannich reaction and the synthetic process of serotonin-avidin conjugate was optimized by controlling the initial molar ratios of serotonin, formaldehyde and avidin (1:12,000:25). The developed bioluminescence immunoassay for serotonin showed good sensitivity (LOD of 0.68 ng/mL) with wide area of dynamic range (5.0×10−10 M~5.0×10−7 M). (cf. the range for serotonin in human blood serum is 151±45 ng/mL). In addition, cross-reactivity studies demonstrated that 5-methoxytryptamine showed some cross-reactivity (28.0%), whereas 3-methylindole, melatonin and 5-hydroxylindole-3-acetic acid showed no crossreactivity,and good recoveries were obtained in serum. Thus, this developed method provides a good tool to monitor serotonin in serum.
Abstract
A sensitive competitive heterogeneous bioluminescence immunoassay for serotonin was developed using photoprotein, aequorin as a label for the first time with the optimal assay conditions; especially, serotoninavidin conjugate was prepared by Mannich reaction and the synthetic process of serotonin-avidin conjugate was optimized by controlling the initial molar ratios of serotonin, formaldehyde and avidin (1:12,000:25). The developed bioluminescence immunoassay for serotonin showed good sensitivity (LOD of 0.68 ng/mL) with wide area of dynamic range (5.0×10−10 M~5.0×10−7 M). (cf. the range for serotonin in human blood serum is 151±45 ng/mL). In addition, cross-reactivity studies demonstrated that 5-methoxytryptamine showed some cross-reactivity (28.0%), whereas 3-methylindole, melatonin and 5-hydroxylindole-3-acetic acid showed no crossreactivity,and good recoveries were obtained in serum. Thus, this developed method provides a good tool to monitor serotonin in serum.
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