Abstract: The immunoassay element for quantitatively analyzing an antigen by determining the change in enzymatic activity of an enzyme-labelled antigen or antibody caused by an immunological reaction. The immunoassay element comprises a substrate layer containing a non-diffusible substrate which forms a diffusible material in the presence of the labelling enzyme, and a reagent layer containing a fragmenting enzyme for further fragmenting the diffusible material into a lower molecular weight product. As the non-diffusible substrate, a substrate capable of reacting solely with the labelling enzyme and incapable of reacting the fragmenting enzyme is utilized. When an endo-active glucosidase is used as the labelling enzyme, and an exo-active glucosidase is used the fragmenting enzyme in the reagent layer, the non-diffusible substrate of the substrate layer is preferred to be an endo type selectively reactive substrate, which means a substrate having a reactivity specific to endo-active glucosidase.

Abstract: A vector includes an enhancer region derived from a transcriptional regulatory region of tyrosine hydroxylase gene wherein the enhancer region enhances transcription amount of a downstream gene in response to a test substance, a promoter which is functionally linked to downstream of the enhancer region, and a reporter gene which is functionally linked to downstream of the promoter.

Abstract: A device for preventing electrical shock from a device with electrical interfaces. A shutter or other barrier associated with the device that physically prevents access or contact to one of the electrical interfaces while another electrical interface is in use.

Abstract: The present invention relates to novel AKT substrate 160 kDa-like protein (AS160-like protein), to a method of identifying a substance altering glucose uptake and/or GLUT4 translocation to the plasma membrane of a cell comprising contacting a test system comprising AKT substrate 160 kDa-like protein (AS160-like protein) with a test substance, and identifying a test substance as a substance altering glucose uptake of a cell by detecting a signal indicative for altered glucose uptake of a cell; a test system comprising a gene coding for the AKT substrate 160 kDa-like protein (AS 160-like protein) and an inducible promoter providing for controllable expression of the gene; the use of the test system for the identification of a substance improving glucose uptake and/or GLUT4 translocation to the plasma membrane of a cell; and the use of AS160-like protein in a model for type 2 diabetes.

Abstract: Compositions and methods for electrochemical detection of an analyte comprising a transition metal compound wherein M is a metallic element that can form a coordinate bond to nitrogen; R and R? are coordinated to M at their nitrogen atoms; L is a linking ligand; Z is chlorine or bromine; m can be from 1 to 6 and X is an anion, or combination of anions, that balances the charge m. Also provided are electrochemical tags and methods of detection.

Abstract: This invention is in the field of medical devices. Specifically, the present invention provides portable medical devices that allow detection of analytes from a biological fluid. The methods and devices are particularly useful for providing point-of-care testing for a variety of medical applications.

Abstract: Substrate probe capable of detecting enzyme activity with high accuracy and a method for detecting the enzyme activity by a multi nuclear magnetic resonance method using the substrate probe. Multi-dimensional nuclear magnetic resonance is performed by using a substrate probe, which is used for measuring enzyme activity by a multi-dimensional nuclear magnetic resonance method and characterized by containing a enzyme recognition site that is selectively recognized by an active-state enzyme, as at least one constitutional unit, and a group to which at least three nuclear magnetic resonance active nuclei each having a nuclear spin and a different resonance frequency are connected, being present specifically to the enzyme recognition, thereby detecting presence of the substrate probe and the enzyme activity. Alternatively, imaging of the enzyme activity is performed by a multi-dimensional nuclear resonance imaging method.

Abstract: A glucose tolerance test device comprising: a test zone having a means to receive at least first and second blood test samples spaced apart by a predetermined time interval; a timer to measure such a time interval, and for example to measure elapsed time after collection of a said first sample; an indicator to indicate at least that a second test is due, operatively Jinked to the timer so as to make such indication after lapse of a predetermined time interval; a data collector to collect data from the lest zone in relation to each test sample; a data register to store data in relation to each test sample, in data communication with said data collector. A method of use is also described.

Abstract: A method is presented for determining the concentration of an analyte in a liquid sample using a reagent system. The reaction of the sample with the reagent system results in a change of the quantity of a fluorophore, the quantity of the fluorophore correlating with the concentration of the analyte, a characteristic measurement variable for the quantity of the fluorophore being measured, and the concentration of the analyte being determined using an evaluation algorithm on the basis of the measurement variable. The reagent system is integrated in an analysis system and the fluorescence lifetime of the fluorophore is used in the ascertainment of the concentration of the analyte.

Abstract: Methods and compositions for modulating glucose transport are provided herein.

Abstract: The invention provides coated sensors for detecting the presence of analytes. The sensor comprises one or more fluorescent sources, such as one or more quantum dots or one or more fluorescent dyes, a polymeric matrix, a surface coating, and one or more analyte sensing components. The surface coating may be a conformal polymeric film, permeable to the analyte, which may be deposited via a solventless process such as initiated chemical vapor deposition or photoinitiated chemical vapor deposition. The surface coating may increase the biocompatibility of the sensor, reduce nonspecific protein adsorption, and/or sequester functional sensor components within the sensor. The invention also provides methods for detecting the presence of an analyte with coated sensors of the invention.

Abstract: A method and apparatus are described that permit an analyte concentration to be estimated from a measurement in the presence of compounds that interfere with the measurement. The method reduces the error in the analyte concentration in the presence of interferents. The method includes the use of a set of measurements obtained for a large population having a range of known analyte and interfering compound concentrations. From a sample measurement, which may or may not be one of the population, likely present interferents are identified, and a calibration vector is calculated.

Abstract: The use of sugar-containing hydrogels as very highly porous, aqueous support material for the immobilization of oligonucleotides, peptides, proteins, antigens, antibodies, polysaccharides, and other biomolecules for sensor applications. Unusually large sizes of interconnected pores allow large target molecules to pass rapidly into and through the gel and bind to immobilized biomolecules. Sugar-containing hydrogels have extremely low non-specific absorption of labeled target molecules, providing low background levels. Some hydrogel materials do not have this type of homogeneous interconnected macroporosity, thus large target molecules cannot readily diffuse through them. Additionally, they nearly always experience non-specific absorption of labeled target molecules, limiting their usefulness in sensor applications. A method is provided for preparing sugar polyacrylate hydrogels with functional chemical groups which covalently bond oligonucleotides and peptides.

Abstract: The use of sugar-containing hydrogels as very highly porous, aqueous support material for the immobilization of oligonucleotides, peptides, proteins, antigens, antibodies, polysaccharides, and other biomolecules for sensor applications. Unusually large sizes of interconnected pores allow large target molecules to pass rapidly into and through the gel and bind to immobilized biomolecules. Sugar-containing hydrogels have extremely low non-specific absorption of labeled target molecules, providing low background levels. Some hydrogel materials do not have this type of homogeneous interconnected macroporosity, thus large target molecules cannot readily diffuse through them. Additionally, they nearly always experience non-specific absorption of labeled target molecules, limiting their usefulness in sensor applications. A method is provided for preparing sugar polyacrylate hydrogels with functional chemical groups which covalently bond oligonucleotides and peptides.

Abstract: Systems and methods for analyzing glucose present in exhaled breath condensate (EBC). In certain embodiments, electrochemical- or coulometnc-based sensing technologies are used to analyze EBC for the presence and/or concentration of glucose. Based on the detected glucose m EBC, the subject invention provides systems and methods for non-invasive, accurate assessment of blood glucose levels.

Abstract: A biosensor system determines analyte concentration from an output signal generated by an oxidation/reduction reaction of the analyte. The biosensor system adjusts a correlation for determining analyte concentrations from output signals at one temperature to determining analyte concentrations from output signals at other temperatures. The temperature-adjusted correlation between analyte concentrations and output signals at a reference temperature may be used to determine analyte concentrations from output signals at a sample temperature.

Abstract: The invention relates to assays for measuring ubiquitin ligase activity and for identifying modulators of ubiquitin ligase enzymes.

Abstract: A health monitoring and diagnostic device (LIFESTREAM cholesterol meter) configured as a self-contained testing and diagnostic unit in a clam-shell type case. One side of the case includes a spring-loaded finger stick and a compartment for carrying one or more packages of disposable items including a test strip, a needle for the finger stick, and an alcohol swipe. The other half of the case includes a test strip reader, a key pad, and a liquid crystal display. The meter reads a test strip carrying a droplet of blood and receives additional diagnostic information from the patient, such as age, gender, weight, and family history of heart disease. Within minutes, the meter displays test results, including total cholesterol levels. The meter also displays additional diagnostic results, such as the patient's “cardiac age,” recommended weight loss, and a cardiac risk assessment. The meter also works in connection with a network-based comprehensive health analysis and reporting system.

Abstract: This invention describes a rapid immunochromatographic assay for measuring the ratio of glycated albumin to total albumin in saliva. Patients with diabetes have elevated levels of glucose in their blood that can react with plasma albumin to form glycated albumin. The amount of glycated albumin formed is directly correlated with the level of plasma glucose that the albumin has been exposed to over a period of time. Saliva albumin is derived from plasma albumin and therefore contains glycated and non-glycated albumin fractions that can be measured. The ratio of glycated albumin to total albumin in saliva will provide an indication of the average amount of protein glycation that occurred over the preceding 2-3 week period. The test is performed using a disposable strip that contains the testing reagents and the results are measured in a measuring instrument that automatically reads, calculates and displays the final result.

Abstract: A porous material containing mesopores with immobilized glucose dehydrogenase is provided. The pore size is at least 10 nm.

Abstract: Methods for covering an opening in an analyte meter are provided. Also provided are methods of determining analyte concentration.

Abstract: The present invention makes it possible to realize a chemical reaction cartridge, which enables prescribed protocols to be achieved easily without differences among operators, which is sealed and disposable, and which has a safe structure against viruses or dangerous drugs. This cartridge is intended to be used for performing chemical reaction of samples, comprising a container formed of a rigid substrate and an elastic body, wherein two or more chambers linked or arranged to be able to be linked through flow paths are formed in said container, and wherein said flow paths, said chambers or both are partially closed by applying an external force to said elastic body from outside said container, thereby being able to move or block fluid substance in said flow paths or chambers.

Abstract: A reagent for blood analysis includes: (1) a compound having the general formula I; and (2) at least one surfactant selected from cationic surfactants and nonionic surfactants. In another aspect a method for differentiating and counting blood cells is provided, the method includes the following steps: (a) mixing a blood sample with the reagent for blood analysis according to the present disclosure to form a cell suspension; (b) detecting scattered light signals and fluorescence signals of cells in the blood sample; and (c) differentiating and counting the cells in the blood sample based upon the scattered light signals and fluorescence signals. The reagent for blood analysis may be effective for identifying and counting erythroblasts and/or basophils in a blood sample to be detected, and meanwhile counting leukocytes therein.

Abstract: The present invention relates to a method for measuring the concentration of a particular component in a blood sample containing blood cells based on a variable correlated with the concentration of the particular component. In the present invention, a concentration (S) in blood plasma obtained by removing blood cell components from the blood sample, a concentration (DI) in the blood sample computed by a differential method and a concentration (EP) in the blood sample computed by an equilibrium point method are expressed by a relational expression which is unrelated to the proportion of the blood cell components in the blood sample, and the concentration of the particular component is computed by using the relational expression.

Abstract: The invention concerns methods and reagent kits for the fluorimetric determination of analytes.

Abstract: It has been found advantageous to deploy coiled antennas as transmitters and receivers for acquiring the dielectric spectrum of materials. This method of impendence spectroscopy has been used to determine the concentration of glucose and other small polar molecules in vitro, as well as in vivo by placement on the antennas so that transmission is through the tissue, as for example on opposite sides of an organ or body part. The optimum selection of antenna coils permits deeper penetration into tissue for glucose detection, improves the SNR as well as expands the spectral range for greater accuracy and precision, to enable continuous non-invasive monitoring for either improved patient or automated management of diabetes.

Abstract: A method for determining an analyte (such a glucose) in a bodily fluid sample includes introducing a bodily fluid sample (e.g., a whole blood sample) into a sample-receiving chamber of an analytical test strip. The method also includes verifying that the bodily fluid sample has filled at least a working portion of the sample-receiving chamber by user visual observation of the working portion through a first portion of a top layer of the analytical test strip, while an opaque second portion of the top layer precludes user visual observation of a non-working portion of the sample-receiving chamber. Thereafter, in the method, the concentration of analyte in the bodily fluid sample is determined only if during the verifying step the user has verified that the bodily fluid sample has filled the working portion.

Abstract: Novel membranes comprising various polymers containing heterocyclic nitrogen groups are described. These membranes are usefully employed in electrochemical sensors, such as amperometric biosensors. More particularly, these membranes effectively regulate a flux of analyte to a measurement electrode in an electrochemical sensor, thereby improving the functioning of the electrochemical sensor over a significant range of analyte concentrations. Electrochemical sensors equipped with such membranes are also described.

Abstract: The present invention relates to a biosensor comprising a substrate with a coating system in which a Ruthenium complex and an enzyme is integrated. The enzyme is able to convert bioproducts, e.g. glucose, fructose or glycerol. The depletion of oxygen during these converting reactions can be monitored via the fluorescence of the Ruthenium complex. The inventive biosensor can be used in biotechnological processes, e.g. the synthesis of biofuels.

Abstract: This disclosure relates to systems, devices, and methods of sensing an analyte. An implantable sensor may be contacted with a test sample under conditions that permit a binding protein and a ligand of the sensor to interact in an analyte-dependent manner to produce an analyte-dependent signal, and (b) detecting the analyte-dependent signal with a detector. A binding protein may reversibly bind an analyte and/or a ligand. A binding protein may have a higher binding affinity for an analyte than for a ligand. A binding protein and a ligand may each include a fluorophore, the absorption and/or emission properties of which may change in an analyte-dependent manner. A binding protein and/or a ligand may be bound to an active or inactive substrate. Some embodiments of systems, devices, and methods may be practiced in vitro, in situ, and/or in vivo. Systems and/or devices of the disclosure may be configured to be wearable.

Abstract: The invention concerns a marker for low-grade inflammation and metabolic syndrome (MS) and MS-related diseases and/or low-grade inflammation-related diseases such as cardiovasculardisease, ischemic heart disease and type 2 diabetes. More particularly it concerns the measurement of the concentration of soluble urokinase plasminogen activator receptor (suPAR) in human biological fluids (sputum, cystic fluid, ascites, serum, plasma, urine) as a tool of diagnosing and/or prognosticating low-grade inflammation and metabolic syndrome and the risk of development of the related diseases such as cancer, cardiovascular disease, ischemic heart disease and type 2 diabetes.

Abstract: Glucose analysis instrument for diabetics, comprising a measuring device for determining glucose concentration values, a displaying device for displaying glucose concentration values, a signaling device for generating a reminder signal, and a control and evaluation device that comprises a processor and a data memory and is used to determine reminder times at which the signaling device is actuated. Event data are stored in the data memory, the event data containing information on events occurring in the life of a user of the glucose analysis instrument and on the time of occurrence of such events. The reminder times are determined by means of a reminder time determination algorithm taking into consideration event data from at least one previous day.

Abstract: A meter and test wand system is capable of cooperatively processing an analysis fluid and communicating results to a user. The system includes a meter case including a case front, a case back, a case top, a case bottom, a first case side member, and a second case side member. The first and second case side members connect the case front to case back. The first case side member has a longitudinal dimension between said case top and said case bottom and a latitudinal dimension between said case front and said case back. The first case side member includes a first case ridge; A test wand is capable of receiving an analysis fluid, and includes a cartridge including a cartridge ridge. The cartridge ridge is sized and shaped to matingly engage with said first case ridge to connect said test wand to said meter case.

Abstract: The present invention provides methods for quantitation of glycated protein in a biological sample using a solid support matrix by making a first bound protein measurement total bound protein under conditions where both glycated and non-glycated protein bind to the support in making a second bound protein measurement under conditions where glycated protein is bound to the support and non-glycated protein is not substantially bound. Diagnostic devices and kits comprising the methods of the present invention are also provided.

Abstract: A sensor utilizing a non-leachable or diffusible redox mediator is described. The sensor includes a sample chamber to hold a sample in electrolytic contact with a working electrode, and in at least some instances, the sensor also contains a non-leachable or a diffusible second electron transfer agent. The sensor and/or the methods used produce a sensor signal in response to the analyte that can be distinguished from a background signal caused by the mediator. The invention can be used to determine the concentration of a biomolecule, such as glucose or lactate, in a biological fluid, such as blood or serum, using techniques such as coulometry, amperometry, and potentiometry. An enzyme capable of catalyzing the electrooxidation or electroreduction of the biomolecule is typically provided as a second electron transfer agent.

Abstract: A method for surface modification of single walled carbon nanotubes. In one embodiment, the method includes the steps of providing a detergent solution, adding a plurality of single walled carbon nanotubes into the detergent solution, performing a first sonication to disperse the single walled carbon nanotubes in the detergent solution, and performing a second sonication after the first sonication to make detergent encased single walled carbon nanotubes. At least one of the plurality of single walled carbon nanotubes is at least partially wrapped by one or more detergent molecules to make it a detergent encased single walled carbon nanotube. In one embodiment, the detergent comprises SDS, PSS or a combination of them.

Abstract: A sensor utilizing a non-leachable or diffusible redox mediator is described. The sensor includes a sample chamber to hold a sample in electrolytic contact with a working electrode, and in at least some instances, the sensor also contains a non-leachable or a diffusible second electron transfer agent. The sensor and/or the methods used produce a sensor signal in response to the analyte that can be distinguished from a background signal caused by the mediator. The invention can be used to determine the concentration of a biomolecule, such as glucose or lactate, in a biological fluid, such as blood or serum, using techniques such as coulometry, amperometry, and potentiometry. An enzyme capable of catalyzing the electrooxidation or electroreduction of the biomolecule is typically provided as a second electron transfer agent.

Abstract: [Problem] In quickly assaying a blood component interfered by glucose and/or its derivative on the bedside or in a clinic or in assaying the same by a patient in his/her own home, there has been required an assay method wherein the whole blood can be used as a sample as such without resorting to a centrifuge or the like. [Means For Solving Problems] A method of assaying a blood component to be used for assaying a blood component interfered by glucose and/or its derivative, characterized by comprising bringing the whole blood into contact with a substance capable of converting glucose and/or its derivative into another substance not interfering the assay and, simultaneously or subsequently, separating blood cells; a device to be used in the assay method; and a kit containing this device.

Abstract: A system and a method for monitoring individual metabolic responses and generating nutritional feedback based on the monitored responses in a qualified subject are disclosed. Measurements of glucose level in the subject are consecutively performed by a measuring device to generate corresponding first data. Subsequently, data is transmitted to an analysis device, where second data representing glycemic responses is generated from a time-series of glucose level measurements represented by the first data. In a next step, the second data is compared with a predetermined individual glycemic response budget for the subject which represents a total amount of individual glycemic responses allowable for a certain time period. Finally, feedback corresponding to the result of the comparison is provided by an output device.

Abstract: A method of manufacturing a microfluidic device having at least one cylindrical microchannel includes providing a substrate, casting an uncured polymer matrix solution onto the substrate, embedding an elongated rod in the uncured polymer matrix solution, curing the polymer matrix solution to form a solidified body, and extracting the elongated rod to form the cylindrical microchannel in the solidified body. In another embodiment, the method includes forming an optical feature on a surface of the microfluidic device. A microfluidic device is also provided, the device including a polymer body, and at least one cylindrical microchannel in the polymer body, the cylindrical microchannel having a diameter between approximately 40 ?m and 250 ?m, inclusive. An additional microfluidic device is provided that functions as an optofluidic spectrometer.

Abstract: A test strip for use of the determination of an analyte in a fluid sample according to one embodiment of the present invention is disclosed. The test strip comprises a base having a top and a bottom, a collection chamber that extends between the top and the bottom of the base, a containing ring that is disposed on the bottom of the base and surrounds the collection chamber, and a capillary channel formed in top of the base that has an inlet fluidly coupled to the collection chamber, a test element disposed within the capillary channel. A lid is attached to the top of the base and covers the collection chamber, the test membrane, and at least a portion of the capillary channel.

Abstract: It is an object of the present invention to provide a means for measuring a concentration of various components rapidly with high accuracy and high sensitivity. The present invention is to provide an oxidative chromogenic compound or salt thereof represented by the following chemical formula (1): wherein R1 and R2 are each independently a linear or branched alkyl group having 1 to 5 carbon atoms; R3, R4, R5, R6 and R7 are each independently a hydrogen atom, a carboxyl group or a sulfonic acid group, or salt thereof, provided that at least one of R3 to R7 is a carboxyl group or a sulfonic acid group, or salt thereof, and the others are hydrogen atoms, and the production method thereof, and the reagent composition and test instrument using the same.

Abstract: Methods and compositions for detection of microbial contaminants in peritoneal dialysis solutions are provided. The methods and compositions employ modified bioburden testing and the detection of peptidoglycan. A novel cause of aseptic peritonitis is provided—aseptic peritonitis associated with gram positive microbial contamination of a dialysis solution. Peptidoglycan is a major component of a gram positive bacterial cell wall and thus can serve as a marker for gram positive bacteria. In this regard, testing for peptidoglycans can be utilized to effectively prevent peritonitis in patients that use the peritoneal dialysis solutions, such as peritoneal dialysis solutions that contain a glucose polymer including an icodextrin and the like.

Abstract: The invention is directed to a method of detecting a biological substance in the nasal secretion and diagnosing a disease following the detection of the biological substance wherein the biological substance is not related to a respiratory disease. The invention also provides treatment of the diseases following the detection of the biological substance and/or diagnosis of the disease. In some embodiments, the diseases are cancer, hepatitis, smell loss, taste loss, diabetes, and leprosy. The invention also provides a kit for diagnosing a disease. The present invention includes methods of analyzing samples from the nose for the detection of biological substances. In particular, nasal secretion or nasal mucus is collected and analyzed for biological substances. The results of this analysis are then suitable for use in diagnosis, prognosis, and determination of suitability of therapeutic interventions.

Abstract: Methods for monitoring and managing glycemia status in a diabetic patient are disclosed. The methods comprise measuring in a diabetic patient, serum concentrations of HbA1c, glucose and inorganic phosphate and comparing the measured concentrations with reference concentrations. An equation representing the reference relationship of HbA1c, glucose and inorganic phosphate is also provided.

Abstract: Described and illustrated herein is an exemplary method of operating an analyte measurement device having a display, user interface, processor, memory, and user interface buttons. Such method can be achieved by measuring an analyte with the analyte measurement device, displaying a value representative of the analyte, querying a user to select a predetermined flag to associate the predetermined flag with the value, and pressing only one of the user interface buttons once to store the predetermined flag with the value in the memory of the analyte measurement device. In one embodiment, the testing device is a glucose meter and the analyte being tested is glucose.

Abstract: A meter adapted for evaluating, saving and simultaneously displaying the number of analyte concentration readings that are greater than a first predetermined analyte concentration level, the number of analyte concentration readings that are less than a second predetermined analyte concentration level and the total number of analyte concentration readings over a predetermined time interval.

Abstract: Provided herein are methods for determining the metabolism of one or more sugars and/or fatty acids, and applications thereof. Such applications include determining the rate of glycogen synthesis and glycolysis, which are believed to be early markers for predicting elevated risk of diabetes and cardiovascular disease. Other applications include methods for screening drugs that effect sugar and/or fatty acid metabolism. The methods are useful for at least partially characterizing drugs for desirable or undesirable (toxic) characteristics. Drugs that are at least partially characterized using the methods of the invention can then be further developed in pre-clinical testing and clinical trials. Such drugs may be found to be useful in treating obesity, diabetes, cardiovascular disease, and other disorders of metabolism.

Abstract: A calibrator suitable for calibrating a noninvasive analyte detector, such as a Raman spectroscopy device, allows the user of the detection device to establish standard measurements of the analyte to be detected, thereby providing assurance that appropriate analyte measurements are obtained at the time of sampling. The calibrator is simple to use, making it suitable for home use by individuals regardless of medical or technical skill level. In particular, the calibrator can be used with a noninvasive glucose detection system, such as for monitoring of blood glucose levels in diabetics. For use with noninvasive detectors designed for obtaining measurements from a subject's fingertip, the calibrator can be formed into a shape that reasonably mimics a fingertip in size, texture and/or spectral properties.

Abstract: An object of the present invention is to provide a more practically advantageous enzyme usable as a reagent for measuring blood glucose than the known enzymes used as blood glucose sensors. A modified flavin adenine dinucleotide dependent glucose dehydrogenase (FADGDH) with more improved heat stability than FADGDH derived from wild-type FADGDH, the modified FADGDH being derived from preferably a eukaryote, more preferably a filamentous fungus, and furthermore preferably an Aspergillus fungus, and, for example, those having a primary structure with at least one amino acid substituted, deleted, inserted or added to FADGDH having an amino acid sequence represented by SEQ ID Nos. 2 or 46 in the sequence table.