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Dimension Vista® System Principles of Vista® Chemistry Photometry/Turbidimetry Online Training

Describe how a photometer measures light absorbance. Define photometric and turbidimetric assay technologies performed. Identify functions of the Dimension Vista System components used during photometric assays. This clinical laboratory training qualifies for continuing education units (CEU).

Define photometric and turbidimetric assay technologies performed by the Dimension Vista® System Describe how a photometer measures light absorbance Identify the functions of Dimension Vista® System components used during photometric assays Welcome to the Photometry online education support tool for the Dimension Vista® System. After completing this course, you’ll be able to: Select Next to continue. 除了鞘液/洗液, 所有试剂都位于分析仪里面。 鞘液/洗液通常放在低于分析仪的地上。 一些试剂包扎起来看作一个整体试剂包。 Congratulations. You’ve completed the Photometry Online Education Support Tool. In this course, you’ve been introduced to the Assay Measurement area of the Dimension Vista® System that performs photometric and turbidimetric assay technologies. Key points of this course are summarized below. Describe how a photometer measures light absorbance Flash lamp power supply: Provides power source to energize xenon gas mixture in flash lamp. Xenon flash lamp: Provides light source to photometer. For every cuvette wheel rotation, the flash lamp emits a beam of light simultaneously to all twelve channels! Fiber optic bundle: Provides 12 channels that guide light to individual detectors. Ten channels are used for photometric measurements, and two channels are used for turbidimetric measurements. Photometer: includes wavelength filters, lenses, a beam splitter, and detectors to measure light intensity readings for each optical channel. Define photometric and turbidimetric assay technologies performed by the Dimension Vista® System Colorimetric assays: Reagent reacts with analyte to produce a color. The photometer measures the amount of light absorbed by the colored solution at a specific wavelength. The amount of light absorbed is directly proportional to the concentration of analyte in the sample. Examples: GLU, CA, BUN, and other tests routinely included in chemistry panels. ACMIA assay: Analyte present in the sample is incubated with enzyme-labeled monovalent antibody to the analyte being measured to ensure that all analyte is bound. Excess monovalent antibody is removed and the mixture is separated magnetically. The supernatant containing the analyte-antibody-enzyme complex is mixed with a substrate and measured using a bichromatic (405 nm and 510 nm) rate technique. The enzyme portion of the antibody-enzyme complex catalyzes the hydrolysis of the substrate causing a change in absorbance at 577nm. The change in absorbance at 577 nm is directly proportional to enzyme activity and is measured using a bichromatic (577, 700 nm) rate technique. Examples: Digoxin (DIG), Digitoxin (DGTX), Cyclosporine (CSA) and Triiodothyronine (T3). EMIT® (Enzyme-Multiplied Immunoassay Technique) assays: Based on competition between analyte present in the sample and analyte in reagent labeled with G6P-DH for binding sites on analyte-specific antibody in the reagent. The amount of free enzyme converts oxidized NAD to NADH, which forms a color. The intensity of color change is proportional to the concentration of analyte in the sample and is measured photometrically at 340 nm. Examples: screens for drugs of abuse, such as barbiturates and ethanol. PETINIA (Particle-Enhanced Turbidimetric Inhibition Immunoassay) assays: Based on competition between analyte present in the sample and analyte in reagent bound to synthetic particles for binding sites on analyte-specific monoclonal antibody in the reagent, which decreases the rate of aggregation. The rate of aggregation is inversely proportional to the concentration of analyte in the sample and is measured using a turbidimetric rate at 340 nm and 700nm. Examples: therapeutic drug monitoring (TDM) tests, such as theophylline (THEO) and tobramycine (TOBR). Identify the functions of Dimension Vista® System components used during photometric assays The photometer is the detection system for both colorimetric and turbidimetric assays that measures and records light absorbance readings. The photometer is located in the Assay Measurement area of the Dimension Vista® System. Dispensing Reagent: A reagent probe aspirates reagent from a Flex® reagent cartridge and dispenses it into a cuvette. Dispensing Sample: The cuvette ring rotates the cuvette into position near the aliquot plate. Sample Probe 1 dispenses sample from an aliquot plate well into the cuvette, where it is mixed with reagent. Incubating Cuvette Contents: The reagent plus sample in the cuvette incubate to allow the chemical reaction to occur. Measuring Light Intensity: After incubation, the cuvette ring rotates to the photometer. As the center of the cuvettes aligns with the center of each wavelength channel, a signal is sent to the photometer, which triggers the xenon lamp to generate one flash to all twelve channels—simultaneously! Clear, parallel surfaces on the cuvettes provide an unobstructed path for the light beam to transmit light through the solution. In addition, the cuvette wheel is positioned on the instrument so that each cuvette aligns with the signal path of each detector. The cuvette ring rotates continuously. At each rotation, the photometer records one reading per cuvette for each wavelength measured. The specific wavelengths measured and the number of measurements recorded depend on the test method. Reusing Cuvettes: After the photometric assay is complete, the cuvette ring rotates to the cuvette washer. Semi-permanent cuvettes are washed and prepared for another assay. Loading Cuvettes: If a cuvette is discarded, the cuvette loader loads a new cuvette into the outer section of the cuvette ring so they are available for testing. Select Next to continue.   Benvenuti al corso Principi di base di Chimica Clinica. La sezione di chimica del laboratorio clinico processa in genere più campioni e referti rispetto a qualsiasi altra sezione del laboratorio. Molti di questi test vengono ordinati dal medico per l'elaborazione immediata, quindi è importante che le persone e gli strumenti lavorino in modo ottimale. La conoscenza dei test eseguiti e delle procedure utilizzate vi darà uno spaccato del funzionamento quotidiano del laboratorio di Chimnica Clinica. In photometric and turbidimetric measurements, reagent and sample are added to a cuvette to produce a substance that absorbs or scatters light at a specific wavelength. Following a predetermined incubation period, the photometer measures the resulting absorbance and converts the measurement to results for reporting. The Dimension Vista® System uses photometric and turbidimetric principles to run a broad menu of routine tests, such as those included in chemistry panels (e.g., GLU, BUN, and CREA). Photometry is also the basis for running special chemistry tests, such as therapeutic drugs and drugs of abuse. What is the difference between turbidimetry and nephelometry? Turbidimetry and Nephelometry Learn about the difference between turbidimetry and nephelometry. Tab TitleTextTurbidimetryIn turbidimetric assays, a photometer is used to measure the reduction of light transmission through a solution or suspension due to particle formation. Turbidimetric assays are ideal for measuring strongly turbid solutions containing many particles or particles that are large compared with the wavelength of visible light.  NephelometryIn nephelometric assays, a nephelometer is used to measure the amount of light scattered toward a detector by the particles in the solution—but the path of scattered light is usually at right angles to the detector. Nephelometric assays are ideal for measuring relatively clear solutions, where the light is only weakly scattered because of a low concentration of particles or the presence of particles that are extremely small compared with the wavelength of the incident light. When complete, select the X in the upper-right corner to close the window and continue. Let’s look inside the Dimension Vista® System to see how the instrument runs photometric assays—from dispensing reagent to obtaining results. In the “virtual laboratory” slideshow, you’ll learn about the following events that occur during photometric assays: Dispensing reagent and sample into cuvettes Aligning cuvettes into the photometer Obtaining photometric readings Reusing semi-permanent cuvettes Key Components of Photometric Measurements Learn about the key components involved in Photometric Measurements. Slide NumberText BlocksCalloutsAudio ScriptImage File1Welcome to the Virtual Chemistry Lab. The photometer is the detection system for both colorimetric and turbidimetric assays that measures and records light absorbance readings. The photometer is located in the Assay Measurement area of the Dimension Vista System. You can see this area of the instrument only in our virtual laboratory. Let's look inside. Select Next to continue.Select the number to review the corresponding text.Note: If audio does not automatically start, select the play arrow in the top left to begin.CalloutsAssay Measurement AreaThe photometer is the detection system for both colorimetric and turbidimetric assays that measures and records light absorbance readings. The photometer is located in the Assay Measurement area of the Dimension Vista® System. You can see this area of the instrument only in our virtual laboratory. Let’s look inside! 2Dispensing Reagent A reagent probe aspirates reagent from a Flex reagent cartridge and dispenses it into a cuvette. Select Next to continue.Select each number to review the corresponding text.CalloutsCuvetteReagent ProbeA reagent probe aspirates reagent from a Flex® reagent cartridge and dispenses it into a cuvette.3Dispensing Sample The cuvette ring rotates the cuvette into position near the aliquot plate. Select Next to continue.Select each number to review the corresponding text.CalloutsAliquot plateCuvette ring The cuvette ring rotates the cuvette into position near the aliquot plate.4Dispensing Sample Sample Probe 1 dispenses sample from an aliquot plate well into the cuvette, where it is mixed with reagent. Select Next to continue.Select the number to review the corresponding text.CalloutsSample Probe 1Sample Probe 1 dispenses sample from an aliquot plate well into the cuvette, where it is mixed with reagent.5Incubating Cuvette Contents The reagent plus sample in the cuvette incubate to allow the chemical reaction to occur. Select Next to continue.The reagent plus sample in the cuvette incubate to allow the chemical reaction to occur.6Measuring Light Intensity After incubation, the cuvette ring rotates to the photometer. Select Next to continue.After incubation, the cuvette ring rotates to the photometer . 7Measuring Light Intensity As the center of the cuvettes aligns with the center of each wavelength channel, a signal is sent to the photometer, which triggers the xenon lamp to generate one flash to all twelve channels-simultaneously! In this way, the photometer provides a dynamic read as the cuvettes move past each channel. Let's take a closer look at the cuvettes. Select Next to continue.Slide QuestionAnswer TextDid you know?The beam of light emitted from the xenon lamp flashes for approximately 1µS to capture the cuvette in the center of the optical window.Select each number to review the corresponding text.CalloutsPhotometerChannelsAs the center of the cuvette aligns with the center of each wavelength channel, a signal is sent to the photometer , which triggers the xenon lamp to generate one flash to all twelve channels simultaneously! In this way, the photometer provides a dynamic read as the cuvettes move past each channel. 8Measuring Light Intensity Clear, parallel surfaces on the cuvettes provide an unobstructed path for the light beam to transmit light through the solution. In addition, the cuvette wheel is positioned on the instrument so that each cuvette aligns with the signal path of each detector. Select Next to continue.Select each number to review the corresponding text.CalloutsCuvetteCuvette RingLight BeamClear, parallel surfaces on the cuvettes provide an unobstructed path for the light beam to transmit light through the solution. In addition, the cuvette wheel is positioned on the instrument so that each cuvette aligns with the signal path of each detector. 9Measuring Light Intensity The cuvette ring rotates continuously. At each rotation, the photometer records one reading per cuvette for each wavelength measured. The specific wavelengths measured and the number of measurements recorded depend on the test method. Select Next to continue.Slide QuestionAnswer TextDid you know?The cuvette ring holds 184 cuvettes and rotates every 3.6 seconds.Select each number to review the corresponding text.CalloutsCuvette RingPhotometerThe cuvette ring rotates continuously. At each rotation, the photometer records one reading per cuvette for each wavelength measured. The specific wavelengths measured and the number of measurements recorded depend on the test method.10Measuring Light Intensity Let's review key components of the photometer: Flash Lamp Power Supply: provides a high voltage power source that energizes the xenon gas mixture in the flash lamp.   Xenon Flash Lamp: provides the light source to the photometer.   Fiber Optic Bundle: distributes light to each of twelve optical channels.   Photometer: includes wavelength filters, lenses, a beam splitter, and detectors to measure light intensity readings for each optical channel. Select Next to continue.Select each number to review the corresponding text.CalloutsFlash Lamp Power SupplyXenon Flash LampFiber Optic BundlePhotometerLet’s review key components of the photometer : The flash lamp power supply provides a high voltage power source that energizes the xenon gas mixture in the flash lamp. The xenon flash lamp provides the light source to the photometer . The fiber optic bundle distributes light to each of twelve optical channels. The photometer includes wavelength filters, lenses, a beam splitter, and detectors to measure light intensity readings for each optical channel. 11Reusing Cuvettes After the photometric assay is complete: The cuvette is either discarded or reused. The cuvette ring rotates to the cuvette washer. Semi-permanent cuvettes are washed and prepared for another assay. Select Next to continue.Select the number to review the corresponding text.CalloutsCuvette WasherWhen the photometric assay is completed, the cuvette is either discarded or re-used, the cuvette ring rotates to the cuvette washer, where the semi-permanent cuvette is washed and prepared for another assay.12Loading Cuvettes If a cuvette is discarded, the cuvette loader loads a new cuvette into the outer section of the cuvette ring so they are available for testing. Select Next to continue.Slide QuestionAnswer TextDid you know?The cuvette loader can load new cuvettes while the Dimension Vista System is processing.Select each number to review the corresponding text.CalloutsCuvetteCuvette LoaderCuvette RingIf a cuvette is discarded, the cuvette loader loads a new cuvette into the outer section of the cuvette ring so they are available for testing. 13You've just learned how photometric assays are performed on the Dimension Vista System, including:   Dispensing reagent and sample into cuvettes Aligning cuvettes into the photometer Obtaining photometric readings Reusing semi-permanent cuvettes When complete, select the X in the upper-right corner to close the window and continue. Congratulations! You’ve just learned how photometric tests are performed on the Dimension Vista® System. Vista Photometric Processing View the entire photometric assay from start to finish.   If the media does not automatically start, select the play arrow to begin. Can you identify what is happening throughout the process? When complete, select the X in the upper-right corner to close the window and continue. The Dimension Vista® System uses “traditional” photometric and turbidimetric assay methods to run routine chemistry tests as well as highly-sensitive techniques to run special chemistry tests. The method used depends on properties of the analyte that needs to be measured.     Photometric Assay Methods Learn about Photometric Assay Methods. Base ImageHotspotsText BlocksImage FilePETINIA PETINIA (Particle-Enhanced Turbidimetric InhibitionImmunoassay) assaysare commonly used to monitor therapeutic drug levels. The assay is based on competition between: Analyte present in the sample, and Analyte in reagent bound to synthetic particles. Both analyte sources compete for binding sites on analyte-specific monoclonal antibody in the reagent, which decreases the rate of aggregation. The rate of aggregation is inversely proportional to the concentration of analyte in the sample and is measured using a turbidimetric rate at 340 nm and 700 nm. Examples of Dimension Vista® System PETINIA assays include therapeutic drug monitoring (TDM) tests, such as theophylline (THEO) and tobramycine (TOBR).  EMIT®  EMIT® assays are commonly used to screen for the presence of drugs. The assay is based on competition between: Analyte present in the sample, and Analyte in reagent labeled with the enzyme glucose-6-phosphate dehydrogenase (G6P-DH). Both analyte sources compete for binding sites on analyte-specific antibody in the reagent. The amount of free enzyme converts oxidized nicotinamide adenine dinucleotide (NAD) to NADH, which forms a color. The intensity of color change is proportional to the concentration of analyte in the sample and is measured photometricallyat 340 nm. Examples of Dimension Vista® System EMIT® assays include screens for drugs of abuse, such as barbiturates and ethanol.    Colorimetry In colorimetric assays, reagent reacts with an unknown quantity of analyte in a sample to produce a color. The photometer measures the amount of light absorbed by the colored solution at a specific wavelength.The amount of light absorbed is directly proportional to the concentration of analyte in the sample. Examples of Dimension Vista® System colorimetric assays include GLU, CA, BUN, and other tests routinely included in chemistry panels.   The ACMIA assay involves the following steps: Analyte present in the sample is incubated with reagent containing an excess of enzyme-labeled monovalent antibody to the analyte being measured to ensure that all analyte is bound. Magnetic particles coated with an analyte analog are added to bind free (unbound) antibody-enzyme conjugate. The reaction mixture is then separated magnetically. Following separation, the supernatant containing the analyte-antibody-enzyme complex is transferred to a cuvette and mixed with a substrate. The enzyme-labeled monovalent antibody that is bound to analyte in the sample is then measured using a bichromatic (405 nm and 510 nm) rate technique. The enzyme portion of the antibody-enzymecomplex catalyzes the hydrolysis of the substrate causing a change in absorbance at 577nm. The change in absorbance at 577 nm is directly proportional to enzyme activity and is measured using a bichromatic (577, 700 nm) rate technique. Examples of Dimension Vista® System ACMIA assays include Digoxin (DIG), Digitoxin (DGTX), Cyclosporine (CSA) and Triiodothyronine (T3).    The Dimension Vista® System photometer provides ten photometric and two turbidimetric channels that guide light toward individual detectors. The optical details of a single channel are shown here. To start the analysis, a sample containing an unknown quantity of analyte to be measured is mixed with reagent. A chemical reaction occurs.   Dimension Vista System Photometer Learn about the photometer on the Dimension Vista System. Base ImageHotspotsText BlocksImage FileA xenon flash lamp provides the light source to the photometer. For every cuvette wheel rotation, the flash lamp emits a beam of light when the center of a cuvette is aligned with the center of a fiber optic channel. The flash of light is simultaneously emitted to all twelve channels.The flash lamp power supply provides a high voltage power source to energize the xenon gas mixture in the flash lamp.The fiber optic bundle provides 12 channels that guide light to individual detectors. Ten channels are used for photometric measurements, and two channels are used for turbidimetric measurements.The reference detector monitors light intensity as a function of time and wavelength.The wavelength filter reduces the light beam to isolate a specific wavelength for each channel. Photometric channels measure light at 293nm, 340nm, 383nm, 405nm, 452nm, 510nm, 540nm, 577nm, 600nm, and 700nm. Turbidimetric channels measure light at 340nm and 700nm.Lenses focus the beam of light directed at the cuvette.The beam splitter directs 10% of the light beam toward a reference detector and the remaining 90% to the cuvette.For photometric and turbidimetric assays, the reaction takes place in semi-permanent cuvettes. As the light beam passes through the cuvette, sample reactants absorb some of the light-the remainder is transmitted.At each cuvette wheel rotation, the measure detector measures the intensity of incoming light and generates an electric signal. The readings are electronically converted to digital signals and sent to the host computer for calculation and analysis. Depending on the test method, the relationship between the intensity of incoming light and concentration of analyte in the sample can be linear or non-linear.