Abbott Laboratories Corporation

Susan E Magic

Jacob J Plattner

Houston F Voss

Steven C March

John W Safford, Jr

Kathryn S Schwenzer

A specific binding assay method employing, as a labeling substance, a reversible trypsin inhibitor for the detection of a hapten. Competition between the hapten to be determined and hapten trysin inhibitor conjugate for antibody to the hapten, in the presence of enzyme, followed by addition of enzyme substrate provides an effective method for hapten analysis. The preferred trypsin inhibitor is a protein having a molecular weight range of 2,000-75,000. The preferred ratio of the hapten to the inhibitor in the conjugate is between 1:1 and 3:1.

Hapten-inhibitor immunoassay

A method for analyzing blood serum or plasma wherein the blood serum or plasma is treated with about 1.4 or more volumes of acetonitrile, propionitrile, tetrahydrofuran or mixtures thereof to precipitate proteins and to provide a supernatant. The separated supernatant is subsequently examined for a component of interest.

Precipitation of protein

The present invention encompasses acid addition salts of a compound of the formula ##STR1## wherein: R.sub.1 represents phenylsulfonyl, benzoyl, carbobenzoxy, and the halo, loweralkyl having 1-4 carbon atoms, loweralkoxy having 1-3 carbon atoms, phenyl, or hydroxy substituted derivatives thereof; or alkanoyl having 2-12 carbon atoms; R.sub.2 represents alkyl having 1-10 carbon atoms, alkoxyalkyl having 2-6 carbon atoms, cycloalkyl having 5-7 carbon atoms, benzyl and the halo, loweralkyl having 1-4 carbon atoms, loweralkoxy having 1-3 carbon atoms, carboxy, hydroxy, or substituted derivatives thereof; R.sub.3 represents hydrogen or guanyl, and m is 3 or 4. Compounds of the present invention are useful as analytical reagents. Enzymatic hydrolysis provides --S--R.sub.2 which can be further reacted with 5,5'-dithiobis (2-nitrobenzoic acid) to provide a colored product by which the enzyme concentration can be determined spectrophotometrically.

Chromogenic substrates

The present invention encompasses the acid addition salts of a compound of the formula ##STR1## wherein R.sub.1 represents phenylsulfonyl, benzoyl, carbobenzoxy, and the halo, loweralkyl having 1-4 carbon atoms, loweralkoxy having 1-3 carbon atoms, phenyl, or hydroxy substituted derivatives thereof; or alkanoyl having 2-12 carbon atoms; and R.sub.2 represents alkyl having 1-10 carbon atoms, or alkoxyalkyl having 2-6 carbon atoms, cycloalkyl having 5-7 carbon atoms, or benzyl and the halo, loweralkyl having 1-4 carbon atoms, loweralkoxy having 1-3 carbon atoms, hydroxy, carboxy, or phenyl substituted derivatives thereof. Compounds of the present invention are useful as analytical reagents. Enzymatic hydrolysis provides --S--R.sub.2 which can be further reacted with 5,5'-dithiobis (2-nitrobenzoic acid) to provide a colored product by which the enzyme concentration can be determined spectrophotometrically.

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Lake Bluff, IL, United States of America

Susan E Magic