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Paper
Analyst, 2009, 134, 1601 - 1607, DOI: 10.1039/b904321g
Potentiometric enzyme immunoassay using miniaturized anion-selective electrodes for detection
Júlia Sz
cs, Ernö Pretsch and Róbert E. Gyurcsányi
An enzyme-linked immunosorbent assay (ELISA) for prostate specific antigen (PSA) detection in human serum was developed based on the potentiometric detection of 6,8-difluoro-4-methylumbelliferone (DiFMU). The assays were carried out in anti-human PSA capture antibody modified microtiter plates (150 µL volume). After incubation in the PSA-containing serum samples,
-galactosidase-labeled PSA tracer antibody was added. The -galactosidase label catalyzed the hydrolysis of 6,8-difluoro-4-methylumbelliferyl--D-galactopyranoside (DiFMUG) and the resulting DiFMU- anion was detected by potentiometric microelectrodes with anion-exchanger membrane. The selectivity of the anion-exchanger electrode is governed by the lipophilicity of the anions in the sample. Since DiFMU- is much more lipophilic (log P = 1.83) than any of the inorganic anions normally present in the working buffers and occurs in its anionic form at the physiological pH (pKa = 4.19), it was chosen as the species to be detected. The potentiometric ELISA-based method detects PSA in serum with a linear concentration range of 0.1–50 ng/mL. These results confirm the applicability of potentiometric detection in diagnostic PSA assays. Owing to simple methodology and low cost, potentiometric immunoassays seem to offer a feasible alternative to the development of in vitro diagnostic platforms.
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