Chemiluminescent and colorimetric aptamer-based assays of human α-thrombinстатья
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Дата последнего поиска статьи во внешних источниках: 20 декабря 2019 г.
Аннотация:Microplate-based sandwich assay for the determination of α-human thrombin (HTb) was developed. Fluorescein-modified 29-mer thrombin binding aptamer (FAM-TBA29) and biotinylated 15-mer thrombin binding aptamer (Bio-TBA15) reacting with different exosites of HTb were used as biorecognition components in the assay. FAM-TBA29 (capture aptamer) was immobilized using its interaction with anti-fluorescein antibody adsorbed on microplate surface. As a sensitive signaling system a combination of Bio-TBA15 and streptavidin-polyperoxidase conjugate was used. Under optimized conditions the detection limit for HTb detection was 1.4 nM; this value was the same in both the colorimetric assay and the chemiluminescent assay. The replacement of colorimetry used for horseradish peroxidase measurement with chemiluminescence increased the assay sensitivity from 0.06 to 1.7 x 106 nM-1 that clearly demonstrated advantage of the latter approach.