Electrochemical Enzyme Immunoassay of Peptide Hormones at pM-Levels

摘要

Electrochemical enzyme immunoassay systems with 10 ng L"' levels were developed for the determination of B- and A-type natriuretic peptides (BNP and ANP), important markers for the diagnosis of heart failure. Sample BNP was added to a solution containing a certain concentration of acetylcholinesterase-labeled anti-BNP antibody, and the enzyme-antibody conjugate that had not reacted with BNP was recovered on a BNP-modified plate. The enzyme activity on the plate was determined with high sensitivity on the basis of the chemisorption/electrochemical desorption process of thiocholine, which was produced through the enzymatic reaction, on a silver electrode. The thiocholine chemisorption (i.e., accumulation) on the silver surface resulted in a sensitivity for the electrochemical determination of enzyme activity that was two orders of magnitude greater than that obtained when using direct measurement without accumulation. The highly-sensitive determination of ANP was carried out by employing alkaline phosphatase-labeied anti-ANP antibody and a glucose oxidase-modified electrode to determine p-aminophenol, which was produced through the phosphatase-catalyzed reaction; based on an enzyme cycling procedure.