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Scheme of ultrasensitive detection of proteins by ELISA coupled with thio-NAD cycling. Our ultrasensitive ELISA requires only the addition of a thio-NAD cycling solution, which includes androsterone derivatives, and 3α-hydroxysteroid dehydrogenase and its coenzymes, to the usual ELISA without any use of special instruments. Absorption (400 nm) of thio-NADH is measured with a commercially available microplate reader.
