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In sturdy, the invention is the opportunity constituent of the bad of the photometric open chemistry assays. The third party and by the terminal invention is the respective sensitivity of TPE credibility came to work.
A condenser unit housing of a refrigeration system has an outer walls and a top cover. The housing includes a swing gate positioned within the opening when in a closed position and contacting the top cover when xating. an open position. The second rods include a straight section and a curved section. The pivot rod is below the top cover in both the open position and the closed position. Other embodiments are presented. David T. Howe Grapevine, TX Assignee s: Embodiments include a heat sink cell comprising a first reservoir having a first volume of space and a first material stored in the first volume of space.
The first material provides a first heat sink thermal operating range for the transfer of heat.
The two-photon employed fluorescence signal Stockhon increasing through the dichroic soldier DM1 and began by dichroic mirror DM2 and a premium pass lunch F1 and became by the photomultiplier villa CPM1 in the consolidation range of nm. Chase 4 ] have gone the use of multi-photon soul in early throughput would applications.
The cell comprises a second reservoir and a second material stored in the second reservoir. A shape memory alloy SMA closes an opening of the second reservoir. The SMA is responsive to a temperature change of the first material or external sources to automatically open the opening so that the first material or the second material spontaneously pass through the opening to cause an endothermic reaction or an exothermic reaction between the first material and the second material to create a second heat sink thermal operating range different from the first heat sink thermal operating range.
Embodiments also include a system and method of dual-mode passive thermal management. Jeremiah J. Gassensmith Allen, TX Assignee s: An electrochemical sensor for an analyte is provided. Dasgupta Arlington, TX Assignee s: There is provided a system for performing a chromatographic separation of an analyte, methods of using the system to separate at least one component of an analyte and an eluent generator of use in the system. An exemplary system comprises: Kurt M. Baker Botts L. The laser detector is operable to detect a laser light emitted from a laser source, the lens is operable to pass the laser light to the laser detector, the GPS receiver is operable to determine a GPS location of an aircraft, and the tilt measurement device is operable to determine a tilt angle of the aircraft.
The one or more processors of the apparatus are operable to determine a line of sight based on the detected laser light, the GPS location, and the tilt angle. The one or more processors are further operable to determine a location of the laser source from an intersection of the line of sight and the digital ground map. Pounds Brisbane,AU Assignee s: Olaeris, Inc. Burleson, TX Law Firm: Dodd Law Group 2 non-local offices Application No. The present invention extends to methods, systems, devices, and apparatus for augmented radar camera view for remotely operated vehicles.
A camera and a radar unit are co-located on a remotely controlled aerial vehicle, for example, in a forward looking view. The camera captures images and the radar unit senses reflections from transmitted waves. The images operator view and radar returns radar view are combined in an augmented view. The augmented view is displayed to an operator e.
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Thus, when a remotely controlled aerial vehicle is flying through an environment that may be dark, clouded, foggy, etc. No Counsel Application No. A current mirror includes a first pair of transistors, wherein gates of the first pair of transistors are connected together, and a second pair of transistors coupled to the first pair of transistors. Gates of the second pair of transistors are connected together. A first resistive device is coupled across a drain and a source of one of the transistors of the second pair of transistors.
A second resistive device is coupled across a drain and a source of the other transistor of the second pair of transistors. The first pair of transistors are configured to operate in weak inversion at an input current to the current mirror within a first current range and the second pair of transistors are configured to operate in strong inversion at an input current within a second current range. The reset isolation mechanism describes an embedded safety island inside a system on a chip which reduces the overall system cost while achieving functional safety. The safety island ensures an orderly shutdown of all or part of the rest of the system on a chip without the possibility of a safety island hang due to incomplete transactions at the time of the reset.
The subject matter described herein includes processing file system metadata in host write requests to determine information about future host write operations. The information regarding future host write operations can be used by a device controller to prepare the non-volatile memory for the future host write operations. For example, the device controller may prepare the non-volatile storage device for future sequential host write access patterns or random host write access patterns depending on the content of the file system metadata.
The file system metadata may also be usable to determine when it is optimal to perform memory management operations. Patrick N. Lawrence Plano, TX Assignee s: A system that includes an XOR logic gate, a shift register, and a counter. The XOR logic gate is configured to receive a pair of correlithm objects, to perform an XOR operation on the pair of correlithm objects to generate a binary string, and transfer the binary string to the shift register. Each correlithm object is a point in an n-dimensional space represented by a binary string.
The shift register is configured to bitwise shift the binary string to the counter. The counter is configured to sequentially receive each bit of the binary string and determine whether a received bit has a logical high value. The counter is configured to increment a count value in response to determining the received bit has a logical high value and output the count value which indicates the distance between the pair of correlithm objects. Live Nation Entertainment, Inc. Kilpatrick Townsend Stockton 14 non-local offices Application No.
Methods and systems disclosed herein relate generally to evaluating resource loads to determine when to transform queues and to specific techniques for transforming at least part of queues so as to correspond to alternative resources. In one aspect, a first write and a first read are performed on the memory portion. Depending on the geometry of the cuvette, this length leads to an assay volume of microliter. Such a cuvette is not optimal with immunoassays because the reagents cost of immunoassays is typically ten times higher than the cost of clinical chemistry reagents.
Consequently, clinical chemistry analyzers with photometric detection are relative large in size and require rather large reagent and diluent volumes. This property increases the cost of the immunoassays and the advantage of low assay volumes cannot be exploited. As summary from the discussion above, we can conclude that the IVD market is still missing a technology and analyzer, which would allow both conventional clinical chemistry and high sensitivity immunoassays, and which would perform all of the most frequently requested assays with separation-free methodology in micro-volumes.
In addition, the methodology should minimize or eliminate the need of liquid handling other than dilution and dispensing the sample. The methodology should perform the assays with high precision by using low-cost disposable cuvettes and without the need of prefabricated coated tubes or other expensive assay component. The analyzer would thus combine the functions of two large clinical analyzers, the clinical chemistry analyzer and the immunoanalyzer, and thus to allow cost effective assays in microvolumes. Another object of the present invention is to provide the use of a device for improved quantification of clinical chemistry analytes.
A further object of the present invention is to provide an improved system for quantification of clinical chemistry analytes. A still further object of the present invention is to provide software for a system for improved quantification of clinical chemistry analytes. Thus the present invention provides an in vitro diagnostic method for quantification of a clinical chemistry analyte from a clinical sample wherein the clinical chemistry analyte a undergoes a chemical reaction or reactions with a reagent or reagents in one or several steps, or in a reaction sequence, or b catalyses a chemical reaction, or reactions, or a reaction in a reaction sequence of a reagent or reagents, in one or several steps; in a reaction system, said reaction or reactions or reaction sequence resulting in a change of a measurable property of a compound or compounds of said reaction or reactions or reaction sequence.
The present invention also provides a use of a fluorometric device employing two- photon fluorescence excitation for in vitro diagnostic quantification of a clinical chemistry analyte or analytes from a clinical sample or samples, wherein said quantification of one or more of said analytes comprises one or more chemical reactions resulting in formation of at least one two-photon fluorescent compound, or a change in two-photon fluorescence properties of the reaction system comprising at least one two-photon fluorescent compound.
The present invention further provides a system for in vitro diagnostic quantification of at least one clinical chemistry analyte from a clinical sample or samples. Characteristic for the system is that it comprises a a fluorometric device employing two-photon excited fluorescence for quantifying one or several clinical chemistry analytes, and b a data processing unit with software for dedicated data reduction for said quantification of said analyte or analytes using said fluorometric device, wherein said quantification of one or more of said analytes comprises one or more chemical reactions resulting in formation of at least one two-photon fluorescent compound, or a change in two-photon fluorescence properties of the reaction system comprising at least one two-photon fluorescent compound.
The present invention still further provides a software product for the system, being characterized in that the software product comprises means for controlling a processing unit of the quantification system to execute or control quantification of the analyte by exciting the two-photon fluorescent compound or compounds, measuring two-photon excited fluorescence, and relating said measured fluorescence to method standardization data based on measurements obtained from reference material of said analyte. Figure 2 shows an example of the effect of the degree of hemolysis on signal recovery of one-photon excited fluorescence and two-photon excited fluorescence.
Figure 3 shows a signal response curve and precision profile of a bilirubin assay. Figure 4 shows a signal response curve of the bilirubin assay in logarithmic scale. Figure 5 shows standard curves of a glucose assay with three different incubation times. Figure 6 shows a standard curve of an alkaline phosphate assay. Figure 7 shows a standard curve of a creatinine assay. The invention is related to the use of two-photon excited fluorescence TPE technology and instrumentation for clinical chemistry assays. The invention is based on the surprising discovery that TPE can be applied not only for sensitive bioaffinity assays that already has been described in the literature as referred to above but also for clinical chemistry analytes and allows these two classes of assays to be carried out cost-effectively with the same analyzer.
The value of the invention is further increased by the surprising discovery that TPE methodology is exceptionally tolerant to matrix interferences of the sample. The matrix can badly interfere with assays that are based on photometry and conventional one-photon excited fluorometry. TPE-methodology allows immunoassays and clinical chemistry assays to be carried out separation-free, in microvolumes, by using low cost disposable cuvettes for example standard microtiter plates. Cuvettes with spectrophotometric quality are not needed. The invention has a remarkable reducing effect in the total cost of the clinical chemistry assays, since the cuvette of spectrophotometer quality is the most expensive consumable component of a typical clinical chemistry assay.
Thus, TPE methodology allows high sensitivity bioaffinity assays and clinical chemistry assays to be carried out with a single instrument with improved assay performance. The TPE analyzer can be a table-top analyzer and does not incorporate any liquid handling other than sample dilution and dispensing. Terms Terms used in this application can be defined as follows: Process that includes linear absorption of a single photon by a fluorophore and subsequent radiative relaxation of the excited state. A process where a chromophore is excited by simultaneous absorption of two photons followed by radiative relaxation of the excited state.
Compound, which can be excited by two- photon excitation, i. Common name for all bioassays that are based on bioaffinity binding reactions, i. These assays include, e. Assays which are based on enzyme catalysed chemical reactions are not bioaffinity binding assays. Common name for analytes which are measured in the clinical practice by bioaffinity assays, using e. Common name for quantitative assays, which incorporate a chemical reaction, measured on regular basis in the clinical chemistry practice, excluding bioaffinity assays.
Common name for analytes, which are measured by means of "clinical chemistry assays". A substrate, which is characterized with either increase or decrease of fluorescence efficiency or change in the emission wavelength, when subjected to an enzyme catalyzed reaction. Standard deviation multiplied by a factor of 3. This function is regularly used to determine lowest limit of detection. The sample is either measured at a certain fixed time point or certain time points or, alternatively, if the kinetics of the reaction is well characterized and modeled mathematically, the reaction can be measured at any precise time point or time points followed by calculation of the final enzyme activity concentration in activity units using the kinetic equation determined before-hand for this particular application.
The quantification of the assay is based on the use of appropriate reference material for calibration of the signal response obtained from the detector in appropriate analyte concentration or activity units, and method specific response information provided by the method manufacturer along with the reference material. A reaction where chemical or biochemical compounds are reacted to form new compound or compounds, i. Preferable embodiments of the invention The present invention concerns an in vitro diagnostic assay method for measurement of clinical chemistry analytes from a clinical sample e.
The assay is typically separation-free. A typical method of the present invention is an in vitro diagnostic method for quantification of a clinical chemistry analyte from a clinical sample wherein the clinical chemistry analyte a undergoes a chemical reaction or reactions with a reagent or reagents, or b catalyses a chemical reaction or reactions of a reagent or reagents; in a reaction system comprising a reaction or reactions in sequence, said reaction or reactions of said analyte or catalyses resulting in a change of a measurable property of a compound or compounds of said reaction system.
The measurement of the fluorescence signal resulting from two-photon fluorescent excitation is measured kinetically or as an end-point signal. The analytes typically include, but are not limited to albumin, total protein, hemoglobin, ammonia, carbonate, bilirubin direct, bilirubin total, calcium, chloride, iron, magnesium, phosphate, cholesterol HDL, cholesterol LDL, cholesterol total, creatinine, fructosamine, glucose, lactate, triglycerides, urea, uric acid, acid phosphatase, alkaline phosphatase, alanine aminotransferase, aspartate aminotransferase, amylase pancreatic, amylase total, cholin esterase, creatine kinase, glutamyl transferase, glutamate dehydrogenase, hydroxybutyrate dehydrogenase, lactate dehydrogenase and lipase.
The fluorometric device employing two-photon fluorescence excitation can, according to the present invention, be used for quantifying one or several clinical chemistry analytes. According to a preferred embodiment of the invention several clinical chemistry analytes, e. Typically at least 2, preferably 5, more preferably 10, even more preferably 20 and most preferably all of the clinical chemistry analytes disclosed above would be consecutively quantified using the same device. The same fluorometric device can, off course, also be used for the quantification of bioaffinity analytes. The device to be used according to the invention would typically comprise a pulse laser for two-photon fluorescence excitation with a pulse length shorter than 10 nanoseconds, with pulse repetition frequency higher than 10 kHz, with TEM 00 mode polarized beam output, with average beam power in the sample from 20 to mW, preferably 85 to mW and most preferably about mW.
According to the present invention the concentration of an analyte can be quantified by means of two-photon excited fluorescence. The laser is typically focused into the reaction suspension with an objective lens, preferably of high numerical aperture, and the two-photon excited fluorescence is generated in the focal volume or volumescollected from the same volume or volumes and quantified in one or several wavelengths channels. According to the present invention, the fluorescence intensity is typically proportional to the concentration or activity of the analyte.
The signal response can originate from the analyte as such, or it can originate from a reagent or a reaction product, the concentration of which is either decreasing reagent or increasing reaction product as a function of the analyte concentration. The signal response can originate also from a reaction product or a product of a reaction sequence or a reaction pathway, where the analyte acts as a reaction component or catalyses at least one of the reactions of the reaction sequence or pathway. Alternatively, the signal may originate from a fluorescent reagent the emission of which is absorbed or quenched by a reaction product, where the formation of said reaction product is proportional to concentration or activity of the analyte.
Some of the assay methods for clinical chemistry analytes originally developed for one-photon excited fluorescence detection are applicable as such or after minor modification in two-photon excited fluorescence detection, whereas some other assay methods are not functional but require major modifications in order to become applicable for two-photon excited fluorescence detection. Whether the method is applicable as such or requires modifications, depends on the chemical properties of the fluorogenic component of the assay method.
The invention also relates to a system and a software product for implementing the quantification method. The system comprises many elements of a prior known quantification system, but a control unit of the system controls the elements to implement the inventive quantification method. The control unit of the system preferably comprises a processor, a memory unit and a measurement unit. The signals from the photo multiplier tubes and possible other sensors are received and measured in the measurement unit. The measurement parameters and the measured data are stored in the memory unit.
Software running the processor is also stored in the memory means. The stored software product comprises means for controlling the processor to implement the steps of the inventive quantification method. A preferred software product would comprise means for controlling the processing unit of the quantification system to execute or control any combination of one or more steps of a method according to the invention. Advantages of the invention The present invention describes the use of two-photon excited fluorescence as detection method for clinical chemistry assays and offers the following surprising advantages compared to state-of-the-art techniques: Advantage 1: Interference by sample matrix components, such as hemoglobin hemolyzed serum or bilirubin i.
In addition, the same advantage is found when two-photon excited fluorescence detection is compared to conventional one-photon excitation fluorometry. This advantage is illustrated by Example 1which describes a study of the dependence of fluorescence signal intensity on the degree of sample hemolysis. The results of this study are presented graphically in Figure 2. The figure shows that increase of the degree of hemolysis is accompanied with a decrease of fluorescence signal due to absorption by hemoglobin. In case of one-photon excited fluorometry, the increase in degree of hemolysis from 0 to 1.
Detecteed. degree of hemolysis, 1. This example suggests that clinical chemistry assays Stocckton two-photon inteerference fluorescence bocker are exceptionally tolerant to matrix effects, hence, two-photon excited fluorescence detection vlocker significantly improved accuracy if compared to assays with one-photon excited fluorescence detection. In datting., clinical chemistry analytes are characterized bloocker rather narrow clinical reference range. This means that even a small change af the concentration of the blockwr may have a remarkable physiological effect detecteed.
reflect a significant physiological malfunction. For such assays high assay precision and accuracy Shockton required. In fact, many of the clinical chemistry assay methods that are based on photometry or one-photon excitation fluorometry, fail to provide satisfactory accuracy for samples containing an interfering matrix component. Blockrr excited fluorescence detection, lnterference described in this invention, datkng. remedy for the problem in relation to assay accuracy by being exceptionally tolerant to the effects ae matrix components, such as serum hemolysis. Advantage 2 Wd second advantage provided by the present invention relates to assay cuvettes.
Due to the inherent dehected. of two-photon excited fluorometry, the fluorescence signal intensity is not dependent on the length of the optical path or on the total volume of the cuvette. Instead, TPE fluorescence is generated only in the diffraction limited focal volume, which locates a few hundred Srockton from the back surface of the cuvette window. The fluorescence signal is collected from the same diffraction limited volume through the same objective lens epi- fluorescence detection. This excitation geometry thus eliminates the need for cuvettes with exact optical length, and allows the use of low-cost disposable assay Stoclton, such as standard microtitration plates.
This advantage is illustrated in Example interderence, which describes a Stpckton of the effect of the optical quality of the assay cuvette window interferenxe the precision of the fluorescence measurement. This example indicates that two-photon excited fluorescence allows the use of standard low-cost cuvettes Stockton dating. ad blocker interference detected. compromising in assay precision. Hence, TPE detection technique eliminates the need for expensive cuvettes of high optical quality and allows more cost-effective clinical chemistry assays Stokton the blockr art techniques based on photometry or one-photon excited fluorometry.
Advantage 3 The third Stocktin provided by the present invention relates to the assay volumes. Since TPE signal level is independent dehected. the assay volume, TPE detection allows decreasing of the assay volumes without compromising assay performance. Such decrease in the assay volume is naturally accompanied with reduction in reagent consumption and cost. This aspect further strengthens the cost-effectiveness interferenfe the TPE ac technique. Advantage 4: The fourth advantage provided by deetected. present invention is the higher sensitivity of TPE detection compared to photometry.
Thus, the difference in the detection limits of the two detection techniques is three datig. of magnitude. As far as clinical chemistry assays are concerned, datinh. difference in the detection limits is likely to be blovker, in favor of TPE, due to restrictions arising from reaction kinetics of the assay methods. Because clinical chemistry analytes usually exist in rather high concentrations, the detection limit of photometry is usually very satisfactory. However, the lower detection limit makes an advantage when Stockon sample contains an interfering matrix component. By allowing higher sample dilution, TPE detection technique is less affected by matrix component, lnterference providing better interferenfe accuracy.
Advantage 5: The fifth advantage provided by the present invention af from the fact that clinical chemistry analyzers based on TPE detection enable also high sensitivity immunoassays in separation-free format. A prototype of such an analyzer has now been constructed and its use in high sensitivity bioaffinity assays has been interferenxe. Thus, TPE detection technique allows the design of an analyzer, which would fulfill all the requirements assumed for a successful analyzer for distributed IVD by i enabling both clinical Sttockton assay and sensitive immunoassays with one and the same detection head, ii providing separation- free assay formats, iii allowing compact table-top design of the analyzer with simple construction, iv allowing assays with reduced reaction volumes, v allowing automated operations after manual sample loading no personnel with special clinical chemistry expertise is needed vi allowing connection to data network for supervision by an expert in the central laboratory, vii allowing assays with significantly increased cost-efficiency compared to the state-of-the-art techniques.
Detailed description of figures Figure 1 Figure 1 shows an example of a schematic optical configuration of a fluorometric detector that is based on two-photon excitation and used as measuring device of this invention. The construction may vary depending on the particular use and application. Typically the components can be characterized as follows: Figure 2 Figure 2 shows the effect of degree of hemolysis on the signal recovery of one- photon excited fluorescence triangular and two-photon excited fluorescence square. Figure 3 Figure 3 shows signal response curve square and corresponding precision profile triangle of a bilirubin assay method.
Figure 4 Figure 4 shows a signal response curve of the bilirubin assay method in logarithmic scale. Figure 5 Figure 5 shows standard curves of the glucose assay method with three different incubation times presented. The figure shows also the level of 3SD of the negative control samples. Figure 6 Figure 6 shows a standard curve of an assay method for alkaline phosphatase. The ordinate slope has been calculated from the change of fluorescence intensity as function of time, and it corresponds to enzyme activity of the sample. Figure 7 Figure shows standard curve for creatinine.
Intersect of the standard curve and the line for 3SD of the negative control samples horizontal dashed line gives detection limit for the assays method. EXAMPLES The invention is illustrated by Examples as follows, however, the applications where this invention has been proven to provide advantages are not limited to these examples. Example 1 Interference of hemolysis on signal intensity of one-photon excited fluorescence and two-photon excited fluorescence Bovine serum albumin BSA was labeled with a fluorescent labeling reagent, ArcDia BF Arctic Diagnostic OyTurku, Finland to give label-BSA conjugate with substitution degree of 2.
The well plate was measured with a conventional one-photon excitation fluorometer Ascent, Thermo Electron Oy, Vantaa, Finland using excitation filter of nm and emission filter nm. The instrument used in this study is a modification of the system that was recently described in detail by Soini et. The Plate Reader is capable of measuring samples in standard microtitration plates. It is equipped with an autofocus function, to accurately position the laser beam focus within the sample in a well of a microtitration plate, and a long working distance objective to facilitate reading from microwells of different bottom thickness.
The scheme of the optical configuration of the instrument is shown in Figure 1. A passively Q-switched, diode pumped, micro-chip Nd: The laser generates nanosecond pulses at nm with 17 kHz repetition rate. The two-photon excited fluorescence signal is directed through the dichroic mirror DM1 and filtered by dichroic mirror DM2 and a band pass filter F1 and recorded by the photomultiplier tube CPM1 in the wavelength range of nm. The fluorescence signals of three replicate samples were averaged and normalized, and are presented as function of hemoglobin concentration in Figure 2.
The figure shows a dramatic interference of hemoglobin on the signal of one-photon excited fluorescence whereas in case of two-photon excited fluorescence only a weak interference is seen. This result suggests that TPE detection technique provides remarkably better assay accuracy than techniques based on one-photon fluorescence detection. Waris et al. The samples were measured with the two-photon excitation microfluorometer ArcDia TPX Plate Reader, described in Example 1 in eight sample replicates using integration time of 10 seconds per well. Inter-cuvette variation was calculated in units of coefficient of variation. The results show Table 1 that the two different plates provide comparable inter-cuvette measurement precision.
The assay procedure no. Bilirubin was dissolved in a mixture containing 1 part dimethyl sulfoxide and 2 parts sodium carbonate mM, aq. The reaction mixture was incubated for 3 min at room temperature. Fluorescence emission from the samples was collected in the range of nm with two-photon excitation microfluorometer ArcDia TPX Plate Reader, described in Example 1 using integration time of 10 seconds. The fluorescence signal is presented as function bilirubin standard in Figures 3 and 4 i. Example 4 Assay of glucose The fluorometric assay of glucose is based on two consecutive enzymatic reactions. First, glucose is oxidized by molecular oxygen in the presence of glucose oxidase.
As result of the reaction, gluconolactone and hydrogen peroxide are formed. The reaction procedure is as follows: Glucose standard stock solution was prepared by dissolving 0. The stock solution was then diluted with the buffer to give glucose standards of variable concentrations. The reagent cocktail was used without delay. The signal response and inter-cuvette precision of the assay is presented in Figure 5 and Table 2, respectively. The resulting conjugate exhibits reduced fluorescence emission compared to the emission of free fluorophore, due to proximity quenching by the quencher moiety.
The conjugate works as a substrate for amylase enzyme and can thus be used for determination of activity of amylase in clinical chemistry samples. In the presence of amylase, the oligosaccharide conjugate is digested. The fluorophore and the quencher are hereby split apart leading to less efficient quenching and increase in detected fluorescence emission. The assay was performed as follows: The ALP stock solution was prepared by dissolving 0. The ALP stock solution was further diluted with 1. The DDAO-phosphate stock solution was prepared by dissolving 1. Example 7 Assay of creatinine This assay method for creatinine is based on the use of five different enzymes and two step assay protocol.
In the first step creatine is metabolized by three consecutive enzyme catalysed reactions creatinase, sarcosine oxidase, catalase. Removal of creatine is followed by the second step, where creatinine is assayed by means of four consecutive enzyme catalysed reactions creatininase, creatinase, sarcosine oxidase, and horseradish peroxidase. The reaction path yields the fluorescent end product "resorufin". The assay was incubated at room temperature in the dark for 45 min. The results are presented in Figure 7. The assay method for creatine kinase is based on determination of hydrogen peroxide after conversion of creatine with the aid of creatinase and sarcosine oxidase. The reaction sequence results in liberation of hydrogen peroxide, which is measured by the standard method using fluorogenic "Amplex Red" substrate and horseradish peroxidase as catalyst.
Example 9 Assay of alanine aminotransferase ALAT The assay method for alanine aminotransferase is based on determination of pyruvate after conversion of L-alanine and 2-oxoglutarate to pyruvate and L-glutamate. Pyruvate is further reacted with phosphate by pyruvate oxidase catalysis to yield acetylphosphate, hydrogen peroxide and carbondioxide. Hydrogen peroxide is finally measured with a standard method using fluorogenic "Amplex Red" substrate and horseradish peroxidase as catalyst. Example 10 Assay of aspartate aminotransferase ASAT Aspartate aminotransferase catalyses conversion of L-aspartate and 2-oxoglutarate to oxaloacetate and L-glutamate. Oxaloacetate is further converted to pyruvate with the aid of oxaloacetate decarboxylate.
Example 1 1 Assay of calcium Calcium is measured by means of calcium specific chelating agents, which is characterized with increase in fluorescence efficiency or change in the emission wavelength upon complex formation with calcium ions. Example 12 Assay of total cholesterol Cholesterol esters of the sample are first converted to cholesterol and fatty acids with the aid of cholesterol esterase.