Dual-cardiac marker capillary waveguide fluoroimmunosensor based on tyramide signal amplification |
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Authors: | Aristeidis E Niotis Christos Mastichiadis Panagiota S Petrou Ion Christofidis Sotirios E Kakabakos Athanasia Siafaka-Kapadai Konstantinos Misiakos |
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Institution: | (1) Immunoassay-Immunosensors Laboratory, Institute of Radioisotopes and Radiodiagnostic Products, N.C.S.R. “Demokritos”, Aghia Paraskevi, Athens, 15310, Greece;(2) Biochemistry Lab, Department of Chemistry, University of Athens, Panepistimiopolis, 15771 Athens, Greece;(3) Institute of Microelectronics, N.C.S.R. “Demokritos”, Aghia Paraskevi, 15310 Athens, Greece; |
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Abstract: | The early diagnosis of acute myocardial infarction requires the determination of several markers in serum shortly after its
incidence. The markers most widely employed are the isoenzyme MB of creatine kinase (CK-MB) and the cardiac troponin I (cTnI).
In the present work, a capillary waveguide fluoroimmunosensor for fast and highly sensitive simultaneous determination of
these markers in serum samples is demonstrated. The dual-analyte immunosensor was realized using glass capillaries internally
modified with an ultrathin poly(dimethylsiloxane) film by creating discrete bands of analyte-specific antibodies. The capillary
was then filled with a mixture of sample and biotinylated detection antibodies followed by reaction with streptavidin–horseradish
peroxidase and incubation with a fluorescently labeled tyramide derivative to accumulate fluorescent labels onto immunoreaction
bands. Upon scanning the capillary with a laser beam, part of the emitted fluorescence is trapped and waveguided through the
capillary wall to a photomultiplier placed on one of its ends. The employment of tyramide signal amplification provided detection
limits of 0.2 and 0.5 ng/mL for cTnI and CK-MB, respectively, in a total assay time of 30 min compared to 0.8 and 0.6 ng/mL
obtained for the corresponding assays when the conventional fluorescent label R-phycoerythrin was used in a 65-min assay.
In addition, the proposed immunosensor provided accurate and repeatable measurements (intra-assay and interassay coefficients
of variation lower than 10%), and the values determined in serum samples were in good agreement with those obtained with commercially
available enzyme immunoassays. Thus, the proposed capillary waveguide fluoroimmunosensor has all the required characteristics
for fast and reliable diagnosis of acute myocardial infarction.
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