Abstract: Vinyl sulfides bearing protected arlyoxy substituents and a process for their preparation are provided. In the process a vinyl sulfide is formed by coupling a ketone compound and a thioester compound with a low valent titanium reagent to form both carbon-carbon bonds in the same step. The vinyl sulfide compounds are useful as intermediates which are convertible into triggerable sulfur-substituted dioxetanes by addition of oxygen to the double bond. The sulfur-substituted dioxetanes additionally are useful for producing chemiluminescence or for further conversion into triggerable alkoxy, alkenyloxy, alkynloxy, aryloxy, aralkyloxy, or acyloxy-substituted dioxetanes. Triggerable dioxetanes are useful for producing chemiluminescence and in detecting activating agents in assays such as immunoassays and nucleic acid hybridization assays.

Abstract: Stable, enzymatically triggered chemiluminescent 1,2-dioxetanes with improved water solubility and storage stability are provided. Dioxetanes further substituted with two or more water-solubilizing groups disposed on the dioxetane structure and an additional fluorine atom or lower alkyl group provide superior performance by eliminating the problem of reagent carryover when used in assays performed on capsule chemistry analytical systems. These dioxetanes display substantially improved stability on storage. Compositions comprising these dioxetanes, a non-polymeric cationic surfactant enhancer and optionally a fluorescer, for providing enhanced chemiluminescence are also provided.

Abstract: Methods of detecting analytes or target species using two enzyme-labeled specific binding partners where the two enzymes function in concert to produce a detectable chemiluminescent signal are disclosed. The methods use a specific binding partner labeled with a hydrolytic enzyme to produce a phenolic enhancer in close proximity to a peroxidase-labeled second specific binding partner. The method is useful to detect and quantitate with improved specificity various biological molecules including antigens and antibodies by the technique of immunoassay, proteins by Western blotting, DNA by Southern blotting, RNA by Northern blotting. The method may also be used to detect DNA mutations and juxtaposed gene segments in chromosomal translocations and particularly to unambiguously identify heterozygous genotypes in a single test.

Abstract: Novel methods and compositions which generate chemiluminescence are provided. The compositions comprise a heterocyclic enol phosphate compound and a dihydroxyaromatic compound in which the two hydroxy groups are separated by an even number of ring carbon atoms. Novel methods and compositions for generating chemiluminescence by reaction with a hydrolytic enzyme are provided as well. The compositions comprise a heterocyclic enol phosphate compound and a protected dihydroxyaromatic compound in which one of the hydroxy groups of the dihydroxyaromatic compound is protected with an enzyme-cleavable group. The novel chemiluminescent compositions are useful in methods for producing chemiluminescence for use in assays of hydrolytic enzymes and enzyme inhibitors and in assays employing labeled specific binding pairs.

Abstract: Stable, enzymatically triggered chemiluminescent 1,2-dioxetanes with improved water solubility are provided. Dioxetanes further substituted with two or more water-solubilizing groups disposed on the dioxetane structure provide superior performance by eliminating the problem of reagent carryover when used in assays performed on capsule chemistry analytical systems. Compositions comprising a dioxetane with two or more water-solubilizing groups, a non-polymeric cationic surfactant enhancer and optionally a fluorescer, for providing enhanced chemiluminescence are also provided.

Abstract: Stable, enzymatically triggered chemiluminescent 1,2-dioxetanes with improved water solubility are provided. Dioxetanes further instituted with two or more water-solubilizing groups disposed on the dioxetane structure provide superior performance by eliminating the problem of reagent carryover when used in assays performed on capsule chemistry analytical systems. Compositions comprising a dioxetane with two or more water-solubilizing groups, a non-polymeric cationic surfactant enhancer and optionally a fluorescer, for providing enhanced chemiluminescence are also provided.

Abstract: A chemiluminescent assay method and compositions are described which use a haloalkoxy group-substituted dioxetane which is deprotected by a hydrolytic enzyme to undergo a chemiluminescent reaction. Chemiluminescent 1,2-dioxetane compounds substituted on the dioxetane ring with a haloalkoxy group which can be triggered by a reagent to generate light are disclosed. Haloalkoxy group-substituted dioxetanes are useful for the detection of triggering agents including enzymes. The enzyme may be present alone or linked to a member of a specific binding pair in an immunoassay, DNA probe assay or other assay where the enzyme is bound to a reporter molecule.

Abstract: Novel methods and compositions which generate chemiluminescence are provided. The compositions comprise a heterocyclic enol phosphate compound and a dihydroxyaromatic compound in which the two hydroxy groups are separated by an even number of ring carbon atoms.Novel methods and compositions for generating chemiluminescence by reaction with a hydrolytic enzyme are provided as well. The compositions comprise a heterocyclic enol phosphate compound and a protected dihydroxyaromatic compound in which one of the hydroxy groups of the dihydroxyaromatic compound is protected with an enzyme-cleavable group.The novel chemiluminescent compositions are useful in methods for producing chemiluminescence for use in assays of hydrolytic enzymes and enzyme inhibitors and in assays employing labeled specific binding pairs.

Abstract: Acridans which are reactable with a peroxidase and peroxide. The acridans are characterized by having an aromatic leaving group ArO which is a di- or polyhalosubstituted phenoxy group. The compounds are useful in assays where one member of a binding pair is linked to the peroxidase and for detecting the peroxidase. The method can also be used to detect hydrogen peroxide.

Abstract: A chemiluminescent assay method, compositions, kits and chemiluminescent acridan compounds are described which use a two-step chemiluminescent reaction process. The reaction involves an acridan compound, preferably a derivative of an N-alkylacridan-9-carboxylic acid, which undergoes a reaction with a peroxide compound, a peroxidase enzyme and an enhancer under conditions of time, temperature and pH which permit the accumulation of an intermediate compound, which is subsequently induced to produce a burst of light by raising the pH. The result is generation of very high intensity light from the reaction. The peroxidase enzyme is present alone or linked to a member of a specific binding pair in an immunoassay, DNA probe assay or other assay where the hydrolytic enzyme is bound to a reporter molecule. The method is particularly amenable to automated assays because of the separation of the incubation and light generating steps.

Abstract: Stable, enzymatically triggered chemiluminescent 1,2-dioxetanes with improved water solubility and storage stability are provided. Dioxetanes further substituted with two or more water-solubilizing groups disposed on the dioxetane structure and an additional fluorine atom or lower alkyl group provide superior performance by eliminating the problem of reagent carryover when used in assays performed on capsule chemistry analytical systems. These dioxetanes display substantially improved stability on storage. Compositions comprising these dioxetanes, a non-polymeric cationic surfactant enhancer and optionally a fluorescer, for providing enhanced chemiluminescence are also provided.

Abstract: Novel alkene intermediates useful in the preparation of 1,2-dioxetanes.

Abstract: A chemiluminescent assay method, compositions and kits are described which use a protected phenolic enhancer compound which is deprotected by a hydrolytic enzyme and then enhances a chemiluminescent reaction. The reaction involves an acridan compound, preferably a derivative of an N-alkylacridan-9-carboxylic acid, which is activated to produce light by a peroxide compound and a peroxidase enzyme in the presence of the deprotected enhancer. The result is enhanced generation of light from the reaction. The hydrolytic enzyme is present alone or linked to a member of a specific binding pair in an immunoassay, DNA probe assay or other assay where the hydrolytic enzyme is bound to a reporter molecule.

Abstract: A chemiluminescent assay method, compositions, kits and chemiluminescent acridan compounds are described which use a two-step chemiluminescent reaction process. The reaction involves an acridan compound, preferably a derivative of an N-alkylacridan-9-carboxylic acid, which undergoes a reaction with a peroxide compound, a peroxidase enzyme and an enhancer under conditions of time, temperature and pH which permit the accumulation of an intermediate compound, which is subsequently induced to produce a burst of light by raising the pH. The result is generation of very high intensity light from the reaction. The peroxidase enzyme is present alone or linked to a member of a specific binding pair in an immunoassay, DNA probe assay or other assay where the hydrolytic enzyme is bound to a reporter molecule. The method is particularly amenable to automated assays because of the separation of the incubation and light generating steps.

Abstract: 4-(3-beta-D-galactopyranosylphenyl)-4-methoxyspiro[1,2-dioxetane-3,2'-adama ntane] which is useful in various chemiluminescent assays.

Abstract: A dicationic surfactant for enhancing the chemiluminescence of chemiluminescent compounds particularly dioxetanes which are triggerable is described. The surfactant has the formulaX.sup.- (R.sub.1).sub.3 A.sup.+ CH.sub.2 -Link-CH.sub.2 A.sup.+ (R.sub.2).sub.3 X.sup.-wherein A is selected from the group consisting of phosphorus and nitrogen atoms separately or together, wherein X is an anionic counterion and R.sub.1 and R.sub.2 are selected from the group consisting of unsubstituted and substituted alkyl and aralkyl groups containing 1 to 20 carbon atoms and R.sub.1 and R.sub.2 can be the same or different, and wherein Link is a carbon chain group selected from the group consisting of dialkylenearyl, aryl, alkylene, alkenylene and alkynylene groups containing 4 to 20 carbon atoms. The surfactants are particularly useful in Western and Southern blot assays.

Abstract: Alkene adamantane compounds useful as intermediates for producing the corresponding 1,2-dioxetanes which are useful in chemiluminescent assays.

Abstract: Improved assay methods utilizing stable, enzymatically triggered chemiluminescent 1,2-dioxetanes with improved water solubility are provided. Assays are performed by a capsule chemistry analytical assay method wherein fluid capsules comprising discrete aqueous segments containing either a chemiluminescent dioxetane reagent or an activating agent separated by an oil-based isolating fluid are flowed through a conduit, and subsequently reacted to produce light.The improvement comprises using a dioxetane substituted with two or more water-solubilizing groups disposed on the dioxetane structure to provide the chemiluminescence. Compositions comprising such a dioxetane, a non-polymeric surfactant enhancer and optionally a fluorescer, provide enhanced chemiluminescence and eliminate the problem of reagent carryover when used in assays performed on capsule chemistry analytical systems.

Abstract: Triggerable dioxetanes with a fluorescent substituent bonded or tethered in the dioxetane so as to produce fluorescence from the group are described. The compounds are of the formula: ##STR1## wherein R.sub.1, R.sub.2 and R.sub.3 are carbon containing groups and wherein one of R.sub.1, R.sub.2 and R.sub.3 is a tethered fluorescent substitutent. The compounds are useful in immunoassays and in probes using enzymes or other chemicals for triggering the dioxetanes to produce light from the fluorescent molecule in the group as a signal.

Abstract: A method and test kit using a hydroxyaryl cyclic diacylhydrazide, a peroxide and a peroxidase enzyme on a blotting membrane for detecting DNA, RNA or proteins (polypeptides) is described. The method and kit provides enhanced chemiluminescence in an assay because of the use of the membrane. The method and test kit is particularly useful in Western, Southern and Northern blotting type assays and DNA sequencing.