Abstract: Novel 1,2-dioxetanes with improved chemiluminescent properties, such as signal intensity, S/N ratio, T1/2, etc. are provided by spiroadamantyl 1,2-dioxetanes, wherein the remaining carbon atom of the ring bears an alkoxy, aryloxy, or arylalkoxy substituent, and either a phenyl or naphthyl ring, this aromatic ring bearing, at the meta position on the phenyl group, or a non-conjugated position on the naphthyl ring, a OX moiety wherein X is an enzyme-cleavable group, which when removed from the dioxetane, leaves the oxyanion which decomposies with chemiluminescence, the aryl ring further bearing an electron active substituent Z. The nature and placement of the Z substituent, at a position not adjacent the point of attachment to the dioxetane ring, strongly influences the properties of the dioxetane. Assays, as well as kits for the performance of those assays, include the dioxetane, an enzyme capable of cleaving the X group, and in certain cases, membranes and chemiluminscent enhancement agents.

Abstract: Enzymatically cleavable chemiluminescent 1,2-dioxetane compounds capable of producing light energy when decomposed, substantially stable at room temperature before a bond by which an enzymatically cleavable labile substituent thereof is intentionally cleaved, are disclosed. These compounds can be represented by the formula: ##STR1## wherein: X and X.sup.1 each represent, individually, hydrogen, a hydroxyl group, a halo substituent, an unsubstituted lower alkyl group, a hydroxy (lower) alkyl group, a halo (lower) alkyl group, a phenyl group, a halophenyl group, an alkoxyphenyl group, a hydroxyalkoxy group, a cyano group or an amide group, with at least one of X and X.sup.1 being other than hydrogen; andR.sub.1 and R.sub.2, individually or together, represent an organic substituent that does not interfere with the production of light when the dioxetane compound is enzymatically cleaved and that satisfies the valence of the dioxetane compound's 4-carbon atom, with the provisos that if R.sup.1 and R.sub.

Abstract: A new and improved polymeric membrane for use in biological assays is provided. A blotting assay employing 1,2-dioxetanes as a source of chemiluminescent employs, as an improved membrane, a polymer comprised of at least one monomer of the formula: ##STR1## The membranes reduce background signal, improve sensitivity and reliability.

Abstract: Probes labeled with 1,2-dioxetane precursors can be employed in a variety of assays. The probes may be nucleic acid, peptide nucleic acid, proteins including enzyme, antibody or antigen, steroid, carbohydrate, drug or non-drug hapten. The probe is provided with a 1,2-dioxetane precursor bound thereto, generally either covalently, or a strong ligand bond. The dioxetane precursor moiety is converted to a bound 1,2-dioxetane by exposure to singlet oxygen. These dioxetane (labels) either spontaneously decompose, or are induced to decompose by an appropriate trigger to release light. The trigger may be a change in pH temperature, or an agent which removes a protective group. Assay formats in which these 1,2-dioxetane labeled probes and referents may be used to include hybridization assays, immuno assays, gel-based assays and Capillary Zone Electrophoresis.

Abstract: Dioxetanes that can be triggered by bases are provided which can be used to detect bases in organic solvents, aqueous preparations and detect gaseous bases in the atmosphere. The dioxetanes can be used to detect the release of bases from various labels, as a means to detect the presence of a base released by physical or natural processes, to calibrate light measuring apparatus and to determine the amount of reducing or oxidizing agent present in the base.

Abstract: Compounds having the formula: ##STR1## wherein T is a polycycloalkylidene group (e.g., adamant-2-ylidene); R is a C.sub.1-20 alkyl, aralkyl or cycloalkyl group; and Y is a fluorescent chromophore (eg., m-phenylene), produced by reacting a compound having the formula: ##STR2## with an R-ylating agent (e.g., R.sub.2 SO.sub.4) in the presence of an alkali metal alkoxide in a polar aprotic solvent. Also, compounds having the formula: ##STR3## are produced by reacting a compound having the formula: ##STR4## wherein X is an electronegative leaving group (e.g., a halogen anion such as chloride ion) in the presence of a Lewis base (e.g., a trialkyl-amine) dissolved in an aprotic organic solvent (e.g., benzene or toluene). Also, compounds having the formula ##STR5## are produced by reacting a compound of the formula ##STR6## with a tetra-O-acylated-O-hexopyranoside halide, then hydrolyzing off the protective acyl groups.

Abstract: A novel synthesis of compound having the formula: ##STR1## wherein T is a stabilizing spiro-linked polycycloalkylidene group, R.sup.3 is a C.sub.1 -C.sub.20 alkyl, aralkyl or heteroatom containing group, Y is an aromatic fluorescent chromophore, and Z is a cleavable group which, when cleaved, induces decomposition of the dioxetane ring and emission of optically detectable light, is disclosed. A tertiary phosphorous acid alkyl ester of the formula:(R.sup.1 O).sub.3 Pwherein R.sup.1 is a lower alkyl group, is reacted with an aryl dialkyl acetal produced by reacting a corresponding aryl aldehyde with an alcohol of the formula:R.sup.3 OHwherein R.sup.

Abstract: A novel synthesis of 1,2-dioxetane phosphate alkali metal salts and novel intermediates employed in this synthesis are disclosed. A hydroxyaryl enol ether alkali metal salt having the formula: ##STR1## wherein T can be an unsubstituted or substituted adamant-2'-ylidene group, R.sup.3 can be a methyl group, Y can be a phenyl group and M.sup.+ can be a sodium cation, is reacted with a phosphorohalidate to give the corresponding enol ether ethylene phosphate, which is then reacted with an alkali metal cyanide to give the corresponding enol ether cyanoethyl phosphate diester alkali metal salt intermediate. Singlet oxygen addition to this enol ether cyanoethyl phosphate diester alkali metal salt intermediate to give the corresponding 1,2-dioxetane cyanoethyl phosphate diester alkali metal salt, followed by .beta.-elimination of the cyanoethyl group using an alkali metal hydroxide or the like, gives the corresponding 1,2-dioxetane phosphate alkali metal salt.

Abstract: A chemiluminescent assays for the determination of the presence or amount of a biopolymer in bound assays using 1,2-dioxetanes in connection with AttoPhos.TM. as chemiluminescent substrates for enzyme-labeled targets or probes is provided. Further disclosed is a kit for conducting a bioassay for the presence or concentration of a biopolymer comprising a) an enzyme complex; b) a 1,2-dioxetane; and c) AttoPhos.TM..

Abstract: A new class of stable dioxetanes bears a polycyclic stabilizing group and aryloxy moiety, the oxygen atom of which is provided with a protective group which can be removed by an enzymatic or chemical trigger admixed with the dioxetane. The polycyclic stabilizing group is preferably spiroadamantane. The group further bears an alkoxy, aryloxy, aralkyloxy or cycloalkyloxymoiety which is partially or completely substituted with halogens, particularly fluorine and chlorine. Proper selection of electron active groups on the stabilizing moiety, the aryl group and the OR group yields enhanced enzyme kinetics, superior light intensity and excellent detection sensitivity in various assays.

Abstract: A compound of the formula ##STR1## where n, Q, and Z are as defined useful in chemiluminescent immunoassays.

Abstract: Chemiluminescent bioassays for the presence or concentration of an analyte in a sample use 1,2-dioxetanes as substrates for the enzyme of an enzyme complex that bind to the analyte. The chemiluminescence obtained from the decomposition of the dioxetane triggered by the enzyme through the formation of the corresponding 1,2-dioxetane oxyanion of the enzyme complex is enhanced by the addition of TBQ as an enhancement agent. Other polymeric quaternary onium salts can be used as enhancement agents in conjunction with enhancement additives which improve the ability of the enhancement agent to form hydrophobic regions in the aqueous sample, in which regions the 1,2-dioxetane oxyanion and its chemiluminescent decomposition products can be sequestered. A kit for performing such assays is also provided.

Abstract: A compound having the formula ##STR1##

Abstract: A compound of the formula ##STR1##

Abstract: A compound of the formula ##STR1##

Abstract: A new and improved polymeric membrane for use in biological assays is provided. A blotting assay employing 1,2-dioxetanes as a source of chemiluminescent employs, as an improved membrane, a polymer comprised of at least one monomer of the formula: ##STR1## The membranes reduce background signal, improve sensitivity and reliability.

Abstract: 1,2-dioxetane compounds bearing a proteolytic enzyme specific amino acid or peptide are provided, which amino acid or peptide can be removed by action of the corresponding protease. When the amino acid or peptide is removed, the 1,2-dioxetane decomposes with chemiluminescence, the generation of light providing a rapid, ultra-sensitive and convenient means for detecting the presence of the protease in the sample being inspected. The amount of light generated, or degree of chemiluminescence, can be correlated with the amount of protease present. Immunoassays, as well as DNA hybridization and DNA probe assays are provided.

Abstract: Spiroadamantyl dioxetanes bearing an alkoxy substituent, and a phenyl substituted on the dioxetane ring can be activated to chemiluminesce if the aromatic substituent bears a meta-substituted moiety designated OX, wherein the X is cleaved by an enzyme with which the dioxetane is permitted to come in contact with. The T.sub.1/2 kinetics of the chemiluminescent reaction, as well as the signal intensity and/or quantum yield of the chemiluminescent reaction, can be altered by addition of a chlorine substituent at position 4 on the phenyl ring. Signal strength can further be enhanced by recognized chemiluminescent enhancers.

Abstract: Chemiluminescent bioassays for the presence or concentration of an analyte in a sample use 1,2-dioxetanes as substrates for the enzyme of an enzyme complex that bind to the analyte. The chemiluminescence obtained from the decomposition of the dioxetane triggered by the enzyme through the formation of the corresponding 1,2-dioxetane oxyanion of the enzyme complex is enhanced by the addition of TBQ as an enhancement agent. Other polymeric quaternary onium salts can be used as enhancement agents in conjunction with enhancement additives which improve the ability of the enhancement agent to form hydrophobic regions in the aqueous sample, in which regions the 1,2-dioxetane oxyanion and its chemiluminescent decomposition products can be sequestered. A kit for performing such assays is also provided.

Abstract: Enzymatically clearable chemiluminescent 1,2-dioxetane compounds capable of producing light energy when decomposed, substantially stable at room temperature before a bond by which an enzymatically clearable labile substituent thereof is intentionally cleaved, are disclosed. These compounds can be represented by the formula: ##STR1## wherein: X and X.sup.1 each represent, individually, hydrogen, a hydroxyl group, a halo substituent, an unsubstituted lower alkyl group, a hydroxy (lower) alkyl group, a halo (lower) alkyl group, a phenyl group, a halophenyl group, an alkoxyphenyl group, a hydroxyalkoxy group, a cyano group or an amide group, with at least one of X and X.sup.1 being other than hydrogen; and R.sub.1 and R.sub.2, individually or together, represent an organic substituent that does not interfere with the production of light when the dioxetane compound is enzymatically cleaved and that satisfies the valence of the dioxetane compound's 4-carbon atom, with the provisos that if R.sub.1 and R.sub.