Abstract: Methods, compositions, kits, and a system are disclosed for detecting one or more analytes in a sample. A mixture comprising the (i) sample, (ii) a first binding reagent comprising a cleavage-inducing moiety and a first binding agent specific for an analyte, and (ii) one or more electrophoretic probes each having a second binding agent is subjected to conditions under which binding of respective binding agents occurs. The interaction between the binding agents and the analyte brings the cleavage-inducing moiety within a proximity effective for cleaving a cleavable linkage tethering an electrophoretic tag to the second binding agent, thereby releasing the tag for electrophoretic separation. Separation of different tags occurs by virtue of their distinct electrophoretic mobilities. After separation, a signal amplification moiety on each tag is activated to generate a signal to indicate the presence of a particular analyte in the sample.

Abstract: We disclose methods of sorting or separating mixtures of living cells (e.g., eukaryotic, prokaryotic, mammalian, pathogenic, bacterial, viral, etc.). We perform our methods by activating cell-selective photophoric labels, which photosensitize and chemically reduce a photosensitive metal compound to form metal grains, particles or crystals. The metal adheres to the cells and forms the basis for sorting or separating different cell types. Photophoric labels may include chemiluminescent agents such as peroxidase enzymes activated with peroxidase substrates capable of luminescence. Photosensitive metal compounds may be present in a light-sensitive matrix or emulsion containing photosensitizable metal compounds, which form metal grains, particles or crystals upon exposure to a developer solution. Developer solutions are formulated to substantially allow living cells to remain viable after exposure to the developing solution.

Abstract: The present invention relates to the use of compounds and to methods for the qualitative and/or quantitative determination of the activity of phosphoamidases or protein phosphoamidases specific for hydrolyzing phosphoamide (P—N) bonds N-phosphorylated basic amino acids like phospholysine, phosphoarginine and phosphohistidine.

Abstract: The present invention provides engineered enzymes generated from protein scaffolds combined with Specificity Determining Regions, the production thereof and the use of said engineered enzymes for research, nutritional care, personal care and industrial purposes.

Abstract: Mammastatin has an approximate molecular weight of 44 kDa in its inactive, non-phosphorylated form. Normal mammary cells express functional phosphorylated forms having approximate molecular weights of 53 kDa and 49 kDa. Metastatic mammary cells either do not express Mammastatin at all, or do not express the 53 kDa or 49 kDa, phosphorylated forms. Mammary cancer cells are inhibited in their growth by the administration of phosphorylated Mammastatin.

Abstract: The present invention concerns a further development and use of biological assays to determine the amount or concentration of an active ingredient present in a sample. The enzyme assay of the present invention determines the amount or concentration of protease inhibitors, including retroviral protease inhibitors such as HIV inhibitors.

Abstract: The present invention relates to methods for differentiating demential diseases comprising measuring the concentration of human lipocalin-type prostaglandin D synthase in a sample of a body fluid collected from a subject and kits for differentiating demential diseases comprising an antibody specific to human lipocalin-type prostaglandin D synthase.

Abstract: The invention is based on the development of several sensitive and simple methods for detecting erythropoietin and the erythropoietin receptor and associated antibodies. The methods and reagents are useful for differential diagnosis in Epo and EpoR related clinical problems. Methods and kits for simultaneous measurement of erythropoietin and erythropoietin receptor in a biological sample are described.

Abstract: The present invention provides for novel reagents whose fluorescence increases in the presence of particular proteases. The reagents comprise a characteristically folded peptide backbone conjugated to two fluorophores such that the fluorophores are located opposite sides of a cleavage site. When the folded peptide is cleaved, as by digestion with a protease, the fluorophores provide a high intensity fluorescent signal at a visible wavelength. Because of their high specificity and their high fluorescence signal in the visible wavelengths, these protease indicators are particularly well suited for detection of protease activity in biological samples, in particular in frozen tissue sections. Thus this invention also provides for methods of detecting protease activity in situ in frozen sections.

Abstract: An assay is disclosed for measuring activity of enzymes, such as kinases, phosphatases, and proteases. Measurements of enzymatic activity are accomplished in a homogenous assay format utilizing a fluorescence quenching technique employing paramagnetic metal ions.

Abstract: High throughput screening assays for the identification of potential therapeutic agents able to modulate the activity of enzymes of fatty acid biosynthesis, especially desaturases and/or elongases, are disclosed along with therapeutic uses of the agents identified by such assays for the prevention and/or treatment of diseases related to fatty acid metabolism. Substrates useful in such assays are also described.

Abstract: Novel synthetic enzyme substrates, enhanced to have improved enzymatic specificity, are disclosed. These synthetic enzyme substrates consist of a substrate peptide that has had its specificity further improved by additional synthetic moieties, selected by combinatorial chemistry techniques, that act to sterically block non-target enzymes. These “steric restrictor” moieties may be labeled to produce a detectable signal upon enzymatic reaction. These novel substrates are particularly useful for improved enzyme substrate microarrays. Specific applications for improved protease substrate microarrays are discussed.

Abstract: Disclosed are compounds of the general formulas: wherein Y is a moiety capable of being cleaved by a hydrolytic enzyme; L is a detectable label; X is a linking group; Z is a halogen; and R is hydrogen, an alkyl, or a halogen. Compounds of this general formula can be used as substrates in an analyte-dependent enzyme activation system, such as that employed in connection with catalyzed reporter depositions. Also disclosed are assays using the compounds, and kits for carrying out the assays.

Abstract: The present invention relates to an assay for determining the activity of a G-Protein-Coupled Receptor specific protein kinase by means of an amplified luminescent proximity homo homogenous assay.

Abstract: The present invention concerns the development of a cell-based assay system having improved sensitivity to HCV NS3 protease activity when compared to known assays, which is useful for screening test compounds capable of modulating (particularly inhibiting) HCV NS3 protease activity. This system provides a first construct comprising a transactivator domain joined downstream of the NS3–5 domains of HCV under the control of a non-cytopathic viral promoter system. A second construct is also provided that comprises a reporter gene under the control of an operator sensitive to the binding of the transactivator. The NS3–5 domains encodes the NS3 polyprotein which comprises: the NS3 protease, followed by the NS4A co-factor, the NS4B and NS5A proteins (including any derivative, variant or fragment thereof), terminated by the NS5B protein (including any derivative, variant or fragment thereof) sufficient to constitute a NS5A/5B cleavage site.

Abstract: The present invention provides methods for identifying agents that modulate a level or an activity of a mitochondrial NAD-dependent deacetylase polypeptide, as well as agents identified by the methods. The invention further provides methods of modulating mitochondrial NAD-dependent deacetylase activity in a cell. The invention further provides methods of modulating mitochondrial function by modulating the activity of mitochondrial NAD-dependent deacetylase.

Abstract: The invention relates to novel methods and compositions for the detection of analytes using the nuclear reorganization energy, ?, of an electron transfer process.

Abstract: Methods, compositions and kits are disclosed for determining one or more target polypeptides in a sample where the target polypeptides have undergone a post-translational modification. A mixture comprising the sample and a first reagent comprising a cleavage-inducing moiety and a first binding agent for a binding site on a target polypeptide is subjected to conditions under which binding of respective binding moieties occurs. The binding site is the result of post-translational modification activity involving the target polypeptide. The method may be employed to determine the target polypeptide itself. In another embodiment the presence and/or amount of the target polypeptide is related to the presence and/or amount and/or activity of an agent such as an enzyme involved in the post-translational modification of the target polypeptide.

Abstract: Methods and compositions for measuring N-dealkylating activity of cytochromes P450 enyzmes are disclosed.

Abstract: The present invention provides filter-based assays for measuring binding of compounds to phosphodiesterases (PDEs). The assay permits stoichiometric binding of compounds to PDEs, thereby providing a highly sensitive measure of a PDE binding and inhibition.

Abstract: The invention provides isolated nucleic acid sequences encoding polypeptides having nucleoside phosphoramidase activity, and methods of screening for nucleoside phosphoramidate compounds that are cleaved by a phosphoramidase or for phosphoramidases that are able to cleave phosphoramidate compounds. The invention also provides methods of delivering a nucleoside monophosphate.

Abstract: The present invention is directed to novel substrates for Hu-Asp. More particularly, the invention provides peptide substrates and fusion polypeptide substrates comprising a ?-secretase cleavage site. Methods and compositions for making and using the peptides are disclosed.

Abstract: The invention relates to a stimulus-responding polymer obtained by polymerizing at least a monomer represented by a general formula (1) and a monomer represented by a general formula (2), and a separation method or concentration method of microorganisms, a purification method, detection method or concentration method of nucleic acids, a separating agent, a separation method of biomaterials and a conversion method of materials, which use this polymer, (in the formula, R11 represents hydrogen atom or methyl group, and R12 represents single bond or a straight or branched alkylene group having from 1 to 5 carbon atoms) (in the formula, R13 represents hydrogen atom or methyl group, and R14 represents hydrogen atom, a straight, branched or cyclic alkyl group, alkoxyl group or alkylamino group having from 1 to 10 carbon atoms, an aryl group or a heterocyclic group).

Abstract: The present invention includes an assay useful for identifying inhibitors of Hepatitis C virus (HCV) activity. Particularly, the present invention is directed to a dual HCV assay useful for high throughput screening that quantifies both the amount of HCV RNA replication inhibitory activity associated with a test compound and the amount of cytotoxicity associated with that test compound. The present invention also includes compounds discovered using this assay, compositions containing such compounds and methods of treating Hepatitis C by the administration of such compounds. The present invention also includes reporter assays using enzymes associated with HCV RNA replication, as well as a cell line having ATTC Accession No. PTA-4583.

Abstract: The present invention relates to compositions and methods for assaying homocysteine (Hcy) and thus related moieties, e.g., S-adenosylhomocysteine (SAH) or adenosine. More particularly, assay methods that employ, mutant SAH hydrolase having binding affinity for Hcy, SAH or adenosine but has attenuated catalytic activity, are provided. The modified enzymes and fusion proteins containing the modified enzymes are also provided.

Abstract: The invention provides compositions and methods of use thereof for labeling peptide and proteins in vitro or in vivo. The methods described herein employ biotin ligase mutants and biotin analogs recognized by such mutants.

Abstract: Provided herein are polypeptides designated CVSP14 polypeptides that exhibit protease activity as a single chain or as an activated two chain form. Methods using the polypeptides to identify compounds that modulate the protease activity thereof are provided. The polypeptides also serve as tumor markers.

Abstract: A peptide substrate is identified by type BoNT/B botulinus toxin, characterized in that it incorporates in its structure a fragment of formula PYA-(Z)-pNF, wherein Z represents one or several amino acids, said fragment being cleavable by said toxin.

Abstract: In this application is described substrates for high-throughput assays of clostridial neurotoxin proteolytic activities. Two types of substrates are described for use in assays for the proteolytic activities of clostridial neurotoxins: (1) modified peptides or proteins that can serve as FRET substrates and (2) modified peptides or proteins that can serve as immobilized substrates. In both types a fluorescent molecules is present in the substrate, eliminating the requirement for the addition of a fluorigenic reagent. The assays described can be readily adapted for use in automated or robotic systems.

Abstract: There is provided a method of identifying a PAR cleaving molecule comprising the steps of: (a) providing an immobilized PAR cleavage peptide linked to a reporter molecule; (b) contacting the immobilized PAR cleavage peptide with a PAR cleaving molecule; and (c) detecting release of the reporter molecule.

Abstract: The present invention provides compositions and methods for the regulation of cytokine signaling through the Tumor Necrosis Factor (TNF) pathway. Specifically, the invention provides a novel gene, polypeptide and related compositions and methods for the regulation of ectodomain shedding. In preferred embodiments, methods and compositions for the regulation of TNF Type-1 Receptor ectodomain shedding are provided. The present invention finds use in therapeutics, diagnostics, and drug screening applications.

Abstract: Provided herein are type II transmembrane serine protease 7 (MTSP7) polypeptides. Zymogen and activated forms of these polypeptides as well as single and two chain forms of the protease domain are also provided. Methods using the polypeptides to identify compounds that modulate the protease activity of an MTSP7 are provided.

Abstract: A chemical composition including a fluorescent polymer and a receptor that is specific for both a target biological agent and a chemical moiety including (a) a recognition element, (b) a tethering element, and (c) a property-altering element is disclosed. Both the fluorescent polymer and the receptor are co-located on a support. When the chemical moiety is bound to the receptor, the property-altering element is sufficiently close to the fluorescent polymer to alter the fluorescence emitted by the polymer. When an analyte sample is introduced, the target biological agent, if present, binds to the receptor, thereby displacing the chemical moiety from the receptor, resulting in an increase of detected fluorescence. Assays for detecting the presence of a target biological agent are also disclosed.

Abstract: Provided herein are type II transmembrane serine protease 10 (MTSP10) polypeptides. Zymogen and activated forms of these polypeptides as well as single and two chain forms of the protease domain are also provided. Methods using the polypeptides to identify compounds that modulate the activity of an MTSP10 are provided.

Abstract: The present invention relates to substantially pure mannan-binding lectin associated serine protease-2 (MASP-2) polypeptides and fragments thereof as well as nucleic acids encoding such polpeptides. Furthermore, the present invention relates to uses of a substantially pure polypeptide comprising amino acid sequences derived from mannan-binding lectin associated serine protease-2 (MASP2) or a functional homologue thereof for the production of a pharmaceutical composition as well as pharmaceutical compositions comprising MASP-2 and/or MASP-2 fragments. In addition the present invention relates to inhibitors of MASP-2 and pharmaceutical compositions comparing such inhibitors. Methods for detecting MASP-2 nucleic acid expression are included in the invention.

Abstract: Provided herein are type II transmembrane serine protease 9 (MTSP9) polypeptides. Zymogen and activated forms of these polypeptides as well as single and two chain forms of the protease domain are also provided. Methods using the polypeptides to identify compounds that modulate the protease activity of an MTSP9 are provided.

Abstract: The invention provides methods and compositions for azide tagging of biomolecules. In one embodiment of the invention, proteins are tagged by metabolic incorporation of prenylated azido-analog substrates. Examples of such analogs are azido farnesyl diphosphate and azido farnesyl alcohol. The azido moiety in the resulting modified proteins provides an affinity tag, which can be chemoselectively captured by an azide-specific conjugation reaction, such as the Staudinger reaction, using a phosphine capture reagent. When the capture agent is biotinylated, the resulting conjugates can be detected and affinity-purified by streptavidin-linked- HRP and streptavidin-conjugated agarose beads, respectively. The invention allows detection and isolation of proteins with high yield, high specificity, and low contamination without harsh treatment of proteins.

Abstract: A bioanalytical method for determining catalases and peroxidases using europium ions.

Abstract: This invention provides methods of identifying ligands that are internalized into a cell. The methods typically involve i) contacting the cell with a reporter non-covalently coupled to a ligand; ii) dissociating the reporter from the ligand and removing dissociated reporter from the surface of the cell; and iii) detecting the reporter within said cell (if any is present) where the presence of the reporter within said cell indicates that the ligand binds to an internalizing receptor and is internalized.

Abstract: A method for measuring the activity of NO-synthase, a corresponding method for identification of NO-synthase modulators, in particular for application in a HTS (High Throughput Screening) and use of the corresponding cation-exchange filter-plate membranes.

Abstract: Methods, compositions and kits are disclosed for determining one or more target polypeptides in a sample where the target polypeptides have undergone phosphorylation. A mixture comprising the sample and a first reagent comprising a cleavage-inducing moiety and an IMAC resin for a binding site on a target polypeptide is subjected to conditions under which binding of respective binding moieties occurs. The binding site is the result of phosphorylation activity involving the target polypeptide. The method may be employed to determine the target polypeptide itself. In another embodiment the presence and/or amount of the target polypeptide is related to the presence and/or amount and/or activity of an agent such as an enzyme involved in phosphorylation of the target polypeptide.

Abstract: In this application is described substrates for high-throughput assays of clostridial neurotoxin proteolytic activities. Two types of substrates are described for use in assays for the proteolytic activities of clostridial neurotoxins: (1) modified peptides or proteins that can serve as FRET substrates and (2) modified peptides or proteins that can serve as immobilized substrates. In both types a fluorescent molecules is present in the substrate, eliminating the requirement for the addition of a fluorigenic reagent. The assays described can be readily adapted for use in automated or robotic systems.

Abstract: Disclosed is a compound of Formula (I), wherein: L is a linking agent; B is a binding agent; X is an atom or group suitable for attaching L to the glycerol chain; and R is a straight chain saturated or unsaturated alkyl group having from 8 to 30 carbon atoms, substituted with M? or M? wherein at least one of M? and/or M? is a detectable label. The compound can be used as a lipase substrate in a solid phase-based assay system, such as a scintillation proximity assay, to detect lipase enzyme activity.

Abstract: The invention relates to novel methods and compositions for the detection of analytes using the nuclear reorganization energy, ?, of an electron transfer process.

Abstract: The present invention relates to determining the prethrombotic state, in particular determining an amount or presence of circulating microparticles and/or stimulated procoagulant cells.

Abstract: Many of the effects of nitric oxide are mediated by the direct modification of cysteine residues resulting in an adduct called a nitrosothiol. A method to detect proteins which contain nitrosothiols involves several steps. Nitrosylated cysteines are converted to tagged cysteines. Tagged proteins can then be detected, for example, by immunoblotting and/or can be purified by affinity chromatography. The method is applicable to the detection of S-nitrosylated proteins in cell lysates following in vitro S-nitrosylation, as well as to the detection of endogenous S-nitrosothiols in selected protein substrates.

Abstract: A diagnostic method for determining MIF protein content in a sample using a direct or an indirect detection technique and wherein MIF is a human MIF protein having a molecular weight of approximately 12.5 kDa.

Abstract: A method to measure protein wherein said method internally controls for factors that affects protein activity such as substrate concentration.

Abstract: Methods for diagnosing and monitoring Alzheimer's Disease in a subject comprising measuring hK6 in a sample from the subject. hK6 may be measured using a reagent that detects or binds to hK6, preferably antibodies reactive with hK6.

Abstract: Articles comprising substantially uniform porous coatings, which may be photopatterned, are provided. The use of such porous coatings increases the surface density of attached compounds within, for example, ligand arrays prepared by methods such as regionally selective solid-phase chemical synthesis. Arrays prepared using the porous coatings may be used within a variety of diagnostic and drug discovery assays.