Beckman Instruments, Inc.

Beckman Coulter, Inc.

Timex Corporation

Xtalite Technology Limited/la Technologie Xtalite Limitee

Chan S Oh

Anthony K Cheng

Julie S Kim

James C Sternberg

Paul C Harris

Gordon Kramer

Gene G Y Shen

Elizabeth K Kearns

Fu-Tai A Chen

Craig W Adams

Cheng F Yan

Josephine M Michael

Thomas S Dobashi

Ramon A Evangelista

Paul Y Hsieh

Stephanie D Yancey

Ezekiel Joseph

Elizabeth K Kearne

The invention provides highly sensitive and rapid homogeneous assays which employ particle-enhanced assay formats in concert with capillary electrophoresis and laser-induced fluorescence (LIF) detection to determine the concentration of an analyte of interest in a sample. Such a determination is made by measuring fluorescent signal(s) (i.e., an electropherogram) produced upon LIF of species present in the reaction mixture that are capable of producing such signals. The method of this invention produces simplified electropherograms by reducing the number of signals that must be separated and subsequently measured, and therefore increases the accuracy of the detection and/or quantification of target analyte concentration in a sample.

Particle based homogeneous assays using capillary electrophoresis with laser-induced fluorescence detection

Synthesis of chlorophenol red glucuronic acid

The separation of enantiomeric pairs of compounds by capillary electrophoresis using highly sulfated &agr;-, &bgr;-, and &ggr;-cyclodextrins as chiral selectors is described herein. These charged cyclodextrins have a higher degree of sulfation and narrower heterogeneity than previous cyclodextrin derivatives. CE analyses using highly sulfated cyclodextrins produce improved resolution of a wide variety of chiral drugs, particulaly neutral compounds and amines. The highly sulfated &agr;-,&bgr;-, and &ggr;-cyclodextrins produce separations which complement each other to further expand the number of neutral and basic drugs that can be resolved by capillary electrophorsis.

Chiral separation of pharmaceutical compounds with charged cyclodextrins using capillary electrophoresis

A microfluidic device having opposed spaced apart sheets of siliceous material, such as glass, the opposed facing sides of each of the sheets having hydrophilic and hydrophobic areas which form therebetween a fluid capillary which is hydrophilic and bounded by hydrophobic areas to form an microfluidic hydrophilic fluid pathway. The preferred device is useful for capillary electrophoresis.

Microfluidic electrophoresis device

A trifunctional conjugate is provided having three chemical moieties attached through a spacer moeity. At least two of the chemical moieties are relatively small molecules, usually less than about 7,000 Daltons in size. The spacer moiety is selected to impart certain steric properties to the conjugate. In one embodiment, the binding of a macromolecular specific binding partner to one of the chemical moieties sterically inhibits the binding of a different macromolecule to another chemical moiety. In another embodiment, the binding of a first chemical moiety to a macromolecule restricts the subsequent binding of a second chemical moiety to a proximate location on the same macromolecule. The three chemical moieties are preferably a nitrophenylazido residue, a phenyl boronic acid residue, and a solid support or a label such as biotin. The spacer is preferably cysteine, lysine, glutamic acid, pyroglutamic acid, S-acetylmercaptosuccinic anhydride or .omega.-carbobenzoxylysine. The conjugate is useful in immunoassays and for targeted labeling of proteins. In an assay, a sample is contacted with the conjugate, a limited quantity of analyte binding partner and an excess of small molecule binding partner. Presence of the analyte is determined by detecting the amount of analyte binding partner diverted away from analyte attached to the spacer of the conjugate.

Analyte assay using a trifunctional conjugate

A method for making a preconjugate which includes immunogenic species of a polymorphic analyte. The method is carried out by reacting an activated binding moiety, and a polymorphic analyte at room temperature for between about 10 hours and about 60 hours. The attaching reaction results in an excess of the preconjugate which includes the immunogenic species of the polymorphic analyte. The preconjugate can be used to make an immunoreactive conjugate useful as a developer antigen in a competitive inhibition immunoassay for the polymorphic analyte.

Method for making a preconjugate

Bidentate reagents for rapidly and quantitatively assaying the concentration of pharmacological agents in biological samples are described. The reagents are used in an immunoassay format for determining the concentration of desired, preselected pharmacological agents, such as benzoylecgonine, cocaine, an opiate, PCP, digoxigenin, acetaminophen, carbamazepine, phenytoin, primidone, theophylline, an aminoglycoside antibiotic, vancomycin, quinidine or a cannabinoid.

Reagents and methods for the rapid and quantitative assay of

An assay for determining the presence of an analyte of interest in a test sample containing an interfering substance that interferes with the assay is disclosed. The assay comprises the steps of forming a reaction mixture by combining in an assay medium (i) an assay system containing components for detecting the analyte of interest, (ii) a test sample containing the analyte and an interfering substance, and (iii) an additional amount of the interfering substance in a quantity sufficient to substantially mask the effect of the interfering substance in the test sample. This reaction mixture is incubated under conditions sufficient to allow complex formation. This resultant complex formation is detected and the presence of the analyte in the test sample is determined.

Method of reducing interferences in assays

A trifunctional conjugate is providing having three chemical moieties attached through a spacer moiety. At least two of the chemical moieties are relatively small molecules, usually less than about 7,000 Daltons in size. The spacer moiety is selected to impart certain steric properties to the conjugate. In one embodiment, the binding of a macromolecular specific binding partner to one of the chemical moieties sterically inhibits the binding of a different macromolecule to another chemical moieties. In another embodiment, the binding of a first chemical moiety to a macromolecule restricts the subsequent binding of a second tridentate member to a proximate location on the same macromolecule. The three chemical moieties are preferably a nitrophenylazido residue, a phenyl boronic acid residue, and a solid support or a label such as biotin. The spacer is preferably cysteine, lysine, glutamic acid, pyroglutamic acid, S-acetylmercaptosuccinic anhydride or .omega.-carbobenzoxylysine. The conjugate is useful in immunoassays and for targeted labeling of proteins.

Trifunctional conjugates

Methods for modulating the rates and dose responses of immunoassays through the incorporation of one or more detergents into the immunoassay reaction are disclosed. The methods are particularly suitable for automated immunoassay formats, especially with formats that use analyte-biotin bidentate reagents. The methods may be used to facilitate the detection of any desired, preselected pharmacological agent.

Detergent-facilitated immunoassay for the rapid and quantitative assay

An initial rate photometric immunoassay method useful for detecting and quantifying analytes in various physiological fluids is disclosed. The method is carried out by combining in a liquid medium an undiluted sample of analyte-containing physiological fluid, such as serum, and an excess of an anti-analyte antibody. A substantially constant initial rate of increase of the liquid medium's turbidity, due to the resulting immunoprecipitation reaction, is measured in real time and compared to a calibration curve prepared from known analyte concentrations to detect and quantify the analyte. Detection and quantification of numerous analytes including, haptens, drugs and proteins, in a wide variety of physiological fluids can be rapidly carried out by the present method.

Initial rate photometric method for immunoassay

A novel heterogenous purification method is disclosed. The method removes a macromolecule from a liquid by allowing the macromolecule to undergo a specific binding affinity reaction with a bidentate conjugate. The method is carried out by immobilizing the bidentate conjugate on a solid phase, contacting the bidentate conjugate with the liquid comprising the macromolecule, and separating the immobilized bidentate conjugate from contact with the liquid, thereby removing the macromolecule from the liquid to obtain either purified macromolecule or a purified liquid.

Method of heterogenous purification using a bidentate conjugate

A novel bidentate conjugate has two different chemical moieties, or bidentate members, attached through an adequate spacer moiety. Each bidentate member acts as a small molecule ligand and is capable of specifically binding to a different macromolecular substance. The bidentate members are relatively small molecules, usually less than about 7,000 Daltons in size. The spacer moiety is selected so that both bidentate members can simultaneously bind to their respective specific binding partners. Where the specific binding partners are multivalent, large complexes can be formed. The formation of these complexes can be inhibited by the presence of an unconjugated monovalent bidentate member, such as free analyte from a test sample. The bidentate is of particular use in turbidimetric or nephelometric inhibition immunoassay procedures.

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Diamond Bar, CA, United States of America

Chan S Oh