(a) According to the structure of the reaction device, automatic biochemical analyzer is mainly divided into flow (FLOW SYSTEM), discrete (DISCRETE SYSTEM) two categories.
1. Flow type refers to the determination of the same items to be tested samples and reagents mixed with the chemical reaction in the same pipe flow process is completed. This is the first generation of automatic biochemical analyzers.
2. Separate type means that the chemical reaction of each sample to be measured and reagent mixing are completed in their own reaction cup. There are several types of branches in it.
(1) Typical discrete automatic biochemical analyzer. This type of instrument is the most widely used.
(2) Centrifugal automatic biochemical analyzer, each sample to be tested is mixed with reagents under centrifugal force in their respective reaction tanks, the chemical reaction is completed and measured. Since mixing, reaction and detection are completed almost simultaneously, it has a high analytical efficiency.
3. Bag-type automatic biochemical analyzer is a reagent bag to replace the reaction cup and colorimetric cup, each sample to be tested in the respective reagent bag reaction and determination.
4. Solid-phase reagent self-customized biochemical analyzer (also known as dry chemical automatic analyzer) is a solid-phase reagent in the film or filter paper and other carriers, each sample to be tested in the corresponding test strips for the reaction and determination. The advantages of this analyzer are that it is quick to operate and easy to carry.
(B) the basic structure of a typical discrete automatic biochemical analyzer
1. Sample (SAMPLE) system
Sample includes calibrators, quality control products and patient samples. The system generally consists of sample loading, transportation and distribution devices.
The common types of sample loading and delivery devices are:
(1) SAMPLE DISK, i.e., the carousel for placing samples has a single or multiple internal and external turns, which are placed individually or in conjunction with the reagent carousel or the reaction carousel, and which rotate in conjunction with the sample dispensing arm during operation. Some use a replacement sample tray, divided into work and standby area, in which to place a number of curved sample rack (SECTOR) for the reprint table, the instrument in the determination of the automatic placement of the replacement, are placed on the sample tray of the sample cup or tube height, diameter and depth of certain requirements, some need a special cup of samples, and some can be used directly with the blood test tube. The number of sample trays to be loaded, as well as the number of calibrators, quality control products, routine samples and emergency samples to be loaded, are generally fixed. These should be selected according to the needs of the job.
(2) drive belt type or rail type feeding that is, the test tube rack (RACK) is not continuous, often 10 a rack, by stepping motor drive conveyor belt, the test tube rack sequentially forward, and then a single rack tube by tube traverse to a fixed position, by the sample distribution arm sampling.
(3)Chain feed sample tube fixed arrangement in the cycle of the drive chain, moving horizontally to the sampling position, some instruments can then clean the tube.
Distribution of the sampling device is mostly composed of syringes, stepper motors or drive pumps, sampling arms and sample probes, etc., ① syringe (SYRINE UNIT). According to the diameter of the syringe and the amount of piston movement distance, quantitative absorption of samples or reagents. Its accuracy determines the precision of sample addition, generally accurate to 1 microliter. When the syringe leaks, the first consideration is whether the probe is clogged, followed by wear and tear of the syringe piston, etc. Some dosing systems use volumetric syringe pumps and CNC pulse stepping motors to improve accuracy. ② The sample probe (PROBE) is connected to the dosing arm to draw sample directly. The probes are equipped with liquid level sensors to prevent probe damage and reduce carry-over contamination. Some have a blockage detection alarm system when the probe samples of blood clots and other substances blocked. The instrument will automatically alarm flushing probe, and skip the current sample, the next sample to add samples. Some also have intelligent anti-collision device encountered obstruction of the probe immediately stop the movement and alarm. Even so, it is still informal operation of the wearing parts. In order to protect the probe, in addition to the need to set up in advance according to the height of the sample container, the minimum liquid level height, etc., the sample container specifications, placement and liquid level height and other setup conditions should not be changed arbitrarily. In some instruments, the sampler and the liquid filler combined together, add the sample and add reagents or diluent a probe to complete at once. (iii) Sample filling arm. Connected to the probe, the movement between the sample cup (reagent bottle) and the reaction cup to complete the sampling and adding samples (add reagents). The way it moves has a certain relationship with the efficiency and working life of the instrument. ④ Valve is used to decide the direction of liquid flow. ⑤ Dilution system. Pre-dilution, post-dilution or doubling of the sample, dilution of the standard stock solution series and so on. The dilution method varies from instrument to instrument, so be careful to recognize it. Reagent system also has a dilution function:
2. Reagent (REAGENT) system generally consists of reagent storage and distribution of liquid addition device.
(1) The reagent compartment is often combined with a reagent carousel. Most instruments have the reagent compartment as a refrigerated room to improve the stability of the on-line reagents.
(2) DISPENSE UNIT. Similar to that of the sample system. , the reagent probe can often be pre-warmed to the reagent, and it is desirable to have a lower starting amount of reagent 2 (R2) probe for the dual reagent system in order to work with reagents of different R1/R2 ratios.
(3) reagent bottle (BOTTLE). There are different shapes and sizes. Such as COBAS MIRA PLUS instrument with 4, 10, 15, 35ML and other specifications, the bottom of the bottle is concave, OLYMPUS AU600 instrument with 30 and 60ML two; Hitachi 7060 instrument with 20, 50, 100ML three and other specifications. Should be based on the workload and reagent specifications. Considering the residual dead volume of reagent bottles and the frequency of replacement, it is reasonable to choose. Uniquely designed card-type reagent box, small volume, anti-evaporation, convenient storage.
(4) supporting reagents often have a bar code, the instrument is equipped with a bar code checking system, the type of reagents, batch number, inventory, expiration date and calibration curves, such as cargo traces, such as BECKMANCX7, etc., to check the verification.
(5) Reagent bottle cap automatic switching system, more conducive to reagent preservation. Some instruments can be added in the operation, the replacement of reagents, and some must be suspended in the state.
3. Barcode (BARCODE) reading system
Generally by the scanning system, signal shaping and decoder three parts. Scanning system with a light source to scan the black bar white space between the bar code symbols due to the bar and space on the light reflection of different, different widths of the bar symbols reflect light of different durations, resulting in different intensity of the reflected light. Received by the photoelectric conversion components and converted into the corresponding intensity of the electrical signal, and finally through the signal shaping, decoded by the decoder. The system automatically identifies the sample rack and sample number identifies the reagents, calibration products and their batch number, expiration date, some and can identify the calibration calibration curve and other information.
Laboratory commonly used barcode types are CODE 39, CODE 128, 2 OF 5 STANDARD, INTERLEAVED2OF 5 and so on. To program your own sample barcodes you need a barcode input device and a barcode reading system to match the barcode. Fully automated tube dispensing cum barcode paste preparation system is available.
4. Reaction system
(1) Reaction plate is loaded with a series of reaction colorimetric cups (CUVETTES), mostly in the form of a carousel. Reaction measurement process according to a fixed program, in the sample arm, liquid arm, stir bar, optical path and cleaning device rotation between. Some instruments in the reaction cup to complete the reaction and then inhaled into the colorimetric cup color, now more common reaction and detection in the same colorimetric cup, more efficient, especially for continuous monitoring method. The colorimetric cup is mostly made of hard quartz glass, hard glass, acrylic plastic without UV absorption, etc., with different service life.DIMENSION series of colorimetric cups are manufactured automatically in the machine and sealed automatically, free of rinsing and pollution. Flow cell type is mainly used in small analyzers. Volume generally dozens of microliters, but the pumping pipeline occupies more reaction liquid, multi-sample continuous use, increasing the chance of cross-contamination.
Peristaltic pump (PUMP). Semi-automatic biochemistry need peristaltic pump pumping reaction solution into the flow colorimetric cell for determination. Requirements for regular calibration of the peristaltic pump, that is, through the suction of quantitative water to test the pump suction volume is accurate. Generally are equipped with pump calibration function.
(2) mixing unit (MIXING UNIT), such as the use of multi-head stirring bar (two-head double-cleaning mixing system). The stirring rod often has Teflon non-stick coating to avoid liquid adhesion.
(3) temperature control device biochemical analyzer through the thermostatic control device to maintain the incubation temperature regulation and constant is also controlled by the computer, the ideal incubation temperature fluctuations should be less than ± 01 ℃. There are three ways to maintain constant temperature. ①Air bath thermostat: that is, there is air between the colorimetric cup and the heater. Air bath thermostat is characterized by convenience, speed, no special materials, but the stability and uniformity is slightly worse than the water bath. Roche (ROCHE) COBAS and 0LYMPUS AU2700 system uses the air bath thermostat mode. ② Water bath circulation type: that is, the colorimetric cup is filled with water around the heater to control the temperature of the water. Water bath constant heat is characterized by a constant temperature, but special preservatives are required to ensure clean water, and the circulating water should be replaced periodically. Hitachi system biochemical analyzers used that is the water bath circulation thermostat. ⑧ thermostatic liquid circulation indirect heating type: the structural principle is in the colorimetric cup around the flow of a special thermostatic liquid (with tasteless, non-polluting, inert, non-evaporating and other characteristics). There is a very small air slit between the colorimetric cup and the thermostatic liquid, and the thermostatic liquid reaches the constant temperature by heating the air of the slit, and its temperature stability is better than that of the dry type, which does not need special maintenance compared with the water bath type circulation type.
5. Wash (WASH) system
Probe and stirring rod using torrent type and other ways of automatic rinsing. Cleaning device generally consists of suction needle, spit liquid needle and wipe brush composition. Cleaning workflow for the suction reaction a suction in an injection of pure water a suction dry a wipe dry. Cleaning liquid has alkaline and acid know two kinds. Generally speaking, after sucking out the reaction liquid, the instrument is first rinsed with alkaline liquid, then rinsed with acidic liquid, and finally rinsed three times with deionized water. The function of the wiping brush is to suck out the water hanging shower on the wall of the cup, and there is a negative suction device inside the brush body. Pay attention to the wiping brush for wear and tear during the use process.
It is worth noting that, for routine rinsing can not remove cross-contamination (CARRY-OVER) of the experiment to be handled specially to reduce cross-contamination or carry contamination. For example, cholesterol determination reagents in the bile acid salt on the serum total bile acid determination of interference, in the elimination of cross-contamination procedures, can be entered into the program, directing the total bile acid is not in the test of cholesterol in the colorimetric cup for the determination of the instrument, if it can not be avoided, the instrument than the colorimetric cup for a special rinse, to prevent the occurrence of cross-contamination.
The water temperature of the rinse water is automatically controlled to a temperature similar to that of the thermostatic reaction tank to ensure a constant temperature of the reaction system and to increase the decontamination power. The use of targeted cleaning after emergency assays appears to be more efficient and economical than the use of a fixed comprehensive cleaning program. Water consumption varies widely between instruments.
Systems such as the ABBOTT AEROSET automated biochemistry analyzer have an automatic wash function (SMART WASH) and optimal specimen sequence selection (OSS). That is, the instrument according to the reagents or samples cross-contamination between the combination of items, automatically change the test order to avoid the mutual influence of the analytical items; indeed can not be avoided, the use of selected special cleaning agent for automatic cleaning.
6. colorimetric system
(1) most of the light source with halogen lamps, working wavelength of 325 ~ 800NM. halogen lamps have a shorter service life, generally only 1,000 ~ L 500 hours. When the luminous intensity of the lamp is not enough, the instrument will automatically alarm, should be replaced in a timely manner, part of the biochemical analyzer is a long-life xenon lamp, 24-hour standby can work for several years, the working wavelength of 285-750NM.
(2) colorimetric cup Automatic biochemical analyzer colorimetric cup is also a reaction cup. Colorimetric cup of light diameter 0.5 ~ 0.7 CM varies, usually quartz or high-quality plastic. Aperture small reagent saving, when the colorimetric cup aperture less than 1 CM, part of the instrument can be automatically corrected to 1 CM. biochemical analyzer colorimetric cup automatic rinsing device in the instrument to complete the colorimetric analysis to do automatic rinsing, suction drying action, colorimetric cup in the automatic check after passing the cycle of continued use. To replace the unqualified colorimetric cup in time. If the quartz colorimetric cup is used, the colorimetric cup should be inspected and cleaned regularly.
(3) monochromator and detector of various types of automatic biochemical analyzers applied to visible ultraviolet absorption spectroscopy, that is, the monitoring of 200-700NM light area of a specific wavelength under the chromophore absorbance changes, supplemented by the microcomputer software system to complete the determination of the calculation. The basis for quantification by visible ultraviolet absorption spectroscopy is the LAMBER-BEER law.
Traditional spectrophotometric determination is generally used before the spectroscopy, that is, in the light source between the lamp and the sample cup first to use filters, prisms or grating spectroscopy, through the adjustable slit, and sample "complementary" monochromatic light, irradiation to the sample cup, and then photocells or phototransistors as a detector, the determination of samples on the monochromatic light absorption (absorbance). Then a photocell or phototube is used as a detector to measure the amount of monochromatic light absorbed by the sample (absorbance).
And most modern biochemical analyzers use post-spectroscopic measurement techniques. After the spectrophotometric measurement: a beam of white light (mixed light) to the sample cup, and then a grating spectroscopy, while a row of light-emitting diodes lined up behind the grating as a detector. The advantage of the rear spectrophotometer is that it does not need to move any parts of the instrument colorimetric system, and can be selected at the same time for the determination of dual-wavelength or multiple wavelengths, which reduces the noise of colorimetry, improves analytical accuracy and reduces the failure rate.
Biochemistry instrument monochromator that is, spectroscopic devices, interference filters and grating spectroscopy two types. Interference filters have inserted and rotatable disk type two kinds. Insertion is the need to use the filter into the filter slot, disk type is equipped with the instrument filters are installed in the disk, rotate to the use of the required filters can be. Interference filters are inexpensive, but easy to become damp and moldy, thus affecting the accuracy of the test results, semi-automatic biochemical analyzers use such filters.
The grating spectroscopy can be divided into holographic reflective grating and etched concave grating two. The former is made after covering a layer of metal film on the glass, there is a certain degree of phase difference is easy to be corroded; the latter is the selected wavelengths fixed engraved on the concave glass, wear-resistant, corrosion-resistant, no phase difference. Automatic biochemical analyzers mostly use grating spectroscopy.
7. Program control system
Computer is the brain of the automatic biochemical analyzer, specimen, reagent injection and identification, bar code recognition, thermostat control, flushing control, results printing, quality control monitoring, instrumentation, such as a variety of faults in the alarm are completed by computer control. Instrument generation over generation, the degree of automation is getting higher and higher, some instruments can even complete part of the daily maintenance program. Automatic biochemical analyzer data processing functions are becoming more and more perfect, such as: the reaction process absorbance, a variety of measurement methods, a variety of calibration methods indoor quality control results of statistics, etc., biochemical instrument can be processed. The computer can also look at the patient's data, the instrument's performance indicators, the instrument's operating status. Automatic biochemistry in the quality control and patient results can also be through the instrument computer and laboratory information system (LIS) interface for network management.
The program controller is the hardware part of the system. It mainly includes:
(1) microprocessor and host computer. Used for each unit of the instrument and the overall control, should have programmed operation, fault diagnosis, a variety of data processing and storage and other powerful functions. Generally according to the needs of the instrument function and computer hardware market mainstream products to configure.
(2) display (CRT MONITOR UNIT). Usually operated with a keyboard, mouse, touch screen and so on.
(3) System and supporting software. More WINDOWS-NT or WINDOWS interface, a full graphical design, multi-menu selection, information guide, fault alarm, help tips, man-machine "dialogue", convenient and intuitive, many instruments have instant response curve display.
(4) Through the RS 232 C data interface and other computers, printers and other equipment to transmit data. With artificial intelligence two-way communication system (HOST QUERY), the instrument can be directly to the main computer to ask the basic information of the patient / sample and test items. Some of them have remote communication and monitoring functions, which can be used to remotely control remote testing and maintenance inspections, and realize network work.
Automated biochemistry analyzers all use program-controlled automatic analysis. Once the analysis program has been determined, the work is as simple as inputting the measurement item or code, the instrument can automatically complete the measurement, calculation and report according to the compiled program. The specific control program varies from instrument to instrument and is generally divided into two types: fixed program and self-programmed program. Fixed program is pre-set by the instrument manufacturer, often with the specified reagents; some cannot be changed, and some can be modified by the user. When it is used together with the supporting reagents, it is convenient to work and the quality is more reliable, but the cost is higher. Self-programming program is flexible and practical, easy to develop new projects, emphasizing the flexibility of the program. For example, batch determination process should be able to insert emergency specimen determination at any time without disrupting the original program; single emergency specimen determination operation is simple, less consumption, flexible pre-dilution or repeated determination.
Two, the general workflow of the instrument
The correct application of biochemical analyzers, just mastered the determination of the technical principle is not enough, but also need to have a sufficient understanding of the specific instrument workflow and determination of the calculation method.
(A) general workflow
The workflow can be examined through the instrument's measurement cycle. Focus on the colorimetric cup blank reading point (CB), sample addition point (S), each reagent addition point (R1, R2, ......), reagent blank reading point (RB), each assay reading point (P), the time interval of each point and the total time of the cycle. Each instrument is generally set at a fixed position on the reaction carousel and at a fixed time in the reaction assay cycle to set the sample reagent and dilution addition level, as well as the assay reading point. For example, the HITACHI 7170 adds sample before PO, RL before PL, R2 between P5 and P6, R3 between P16 and P17, and R4 between: P33 and P34 when the cycle from P1 to P34 is 10 minutes.
(ii) Data processing calculation method
The absorbance data read by the instrument at each absorbance reading point are not always included in the concentration calculation. Instruments are often based on the definition of the instrument and the operator to set the requirements of the original absorbance data for the calculation process, converted to the so-called reaction data, and then according to the coefficients or formulas for concentration calculation. Examples are as follows:
1. HITACH 7170 endpoint (A ) absorbance is calculated as (AX + AX-1)/2. Actual absorbance = absorbance data × 10 000.
2. 0LYHPUS AU 600 reagent blank data: P0 point reagent blank (RB) = P0 point absorbance of one. water blank (WB) of the colorimetric cup; reagent blank (RB ) of any measurement point. The absorbance at this point is a colorimetric cup water blank (WB).
3. Response data from the MONARCH 1 000 two-point endpoint method. (absorbance of the endpoint measurement one absorbance of the endpoint blank) one (absorbance of the beginning point measurement one absorbance of the beginning point blank).
4. Response data for the endpoint method (with reagent blank, END method) for AU 600 = endpoint absorbance one absorbance at point P0 (reagent blank, RB). Reaction data for the endpoint method (without reagent blank, END). Method) reaction data = endpoint absorbance - colorimetric cup water blank (WB).
5. Response data for the two-point method (own blank) for AU 600 = [point reading after addition of second reagent a P0 point reading] a [point reading before addition of second reagent a PO point reading].
Three, the main operating procedures
(a) the instrument before the operation of the operating procedures
Mainly for the basic setup of the instrument:
L. test setup on the name of the test, the code, the test combination (PROFILE), the test rounds (ROUND), including the order of the test and other settings, if necessary.
2. Parameter settings for each test including the ratio between the test, the results of the verification and other parameters.
3. agent settings according to the relevant test parameters, set the test reagent bit, reagent bottle specifications, if necessary, set the reagent lot number, expiration date.
4. Calibration product settings for the location of the calibration product, concentration and quantity settings.
5. Quality control settings According to the quality control requirements. Set the number of quality control substances, quality control rules, quality control items and the corresponding quality control parameters.
6. Sample Tube Settings Including the type of sample tube, residual liquid height (dead volume), identification methods and other settings.
7. Other settings include data transmission mode, result report format, review mode and review criteria.
(2) Routine operating procedures
Power on (warm-up, maintenance) - set the start conditions (date and time index, rounds, sample start number, etc.) - as needed, apply for calibration, quality control and patient determination of the project (including rack number, cup number or sequence number. The request can be continued during the assay)_Load calibrators, QC and patient specimens-Load reagents-Check the starting status of the instrument (in particular, check that the assay start number corresponds to the rack number and the request number if the bar code system is not used and the sample is identified sequentially) -Calibrate and QC assays-Check calibration and QC results -Patient specimen assays-Monitor the assay process (reagent checking, observing analytical results, editing, correcting) calibration) - Data transfer (printing of reports, transmission to the test management system, including workload statistics, financial statistics, patient status tracking, QC analysis, etc.). Post-measurement maintenance.
(C) the results of the inspection and analysis
1. To understand and be familiar with the instrument's various warning symbols of the meaning and role of the correct setting of parameters under the premise of the use of a variety of warning symbols can improve the efficiency of their discovery and problem solving.
2. To be familiar with and flexible use of the instrument's related operation screen (interface), such as: the reaction process monitoring (REACTION MONITOR), observe the reaction time process curve; calibration tracking (CALIBRATION TRACE) to review and analyze the calibration curve; the use of statistics (STATISTICS) to understand the average value of the patient's measurements and data analysis of different date periods; the use of analysis data editing; the use of data editing and data analysis; the use of analysis data editing and the use of data editing. Data analysis; the use of analysis data editing (DATA EDIT) to view and correct measurement data.
3. The calibration check should make full use of the function of the instrument settings to monitor the calibration curve graph, the absorbance value of each calibration point (can not ignore the reagent blank value and blank rate value), the calculated K value and other fluctuations, as well as compared with the previous. If necessary, the reaction time course curve should be further examined. The experimental conditions must be grasped in conjunction with the quality control data.
4. Patient results of the examination in addition to visual observation or serum index to understand the specimen traits, attention and understanding of clinical information and diagnosis, learn to analyze the reaction time process curve and data is an important basic skills.
Four, the basic measurement method
(a) end point method (END POINT METHOD)
According to the reaction to reach equilibrium when the reaction products of the absorption spectral characteristics and its absorbance magnitude, the method of quantitative analysis of substances. For general chemical reactions, the end point is when the reaction is complete (or the dynamic equilibrium of positive and negative reactions) and the reaction products are stable. For an antigen-antibody reaction, the end point is when the antigen and antibody have completely reacted to form the largest and stable immune complex. The reaction time course curve is parallel to the X-axis. In the measurement of the calculation method, generally divided into one-point method and two-point method.
1. One point method (ONE POINT) The absorbance value of air blank (GB), water blank (WB) or reagent blank (RB) before mixing the reagents and samples is taken as the base point of measurement, and the absorbance reading at the end point of the reaction is subtracted from that of the blank to get the absorbance of the reaction. The result is obtained by comparing the absorbance of the reaction with that of the calibration solution under the same conditions. It is often used in conjunction with the one-point calibration method, i.e., a calibration concentration is used and the calibration curve passes through the zero point and is linear. It is also used for multi-point calibration.
2. TWO POINT END method (TWO POINT END) that is, the end point of a starting point method to mix the reagents and samples at a certain time point after the starting point, to the end of the reaction of the absorbance readings minus the starting point readings. Under certain conditions, it can reduce the specificity of the sample to the reaction or the reaction itself in the interference (mainly refers to chromaticity interference). Often use two reagents, mostly to add R2 before a point as the starting point of the measurement; in some cases, you can also add R2 after a point as the starting point of the measurement. If you use a single reagent, it is difficult to use when the main reaction is started too quickly or when the starting point of the instrument reading is limited.
Fixed time method (FIXED METHOD) and the two-point endpoint method is only in the difference: the end point of the measurement reading is not in the reaction equilibrium section, but according to the methodology to choose. Such as serum creatinine (picric acid method) determination.
3. Three-point endpoint method, i.e., double endpoint method, in a channel at a time to carry out two reaction-related endpoint method of determination. For example, free fatty acids and triglycerides are measured at the same time. Some instruments (such as the HITACHI series) are set up for this method.
(II) CONTINUOUS MONITORING METHOD
Also known as the rate method (RATE ASSAY). That is, continuous monitoring of the reaction process, based on the measured rate of product generation or substrate consumption quantitative analysis of the method. In the reaction time course curve for the reaction is a constant velocity section (slope remains constant), commonly used in enzyme activity linear reaction period determination.
1. Continuous monitoring method, i.e., zero-level reaction rate method, also known as slope method. In the longer reaction time interval (at least 90-120 seconds), every certain time (often 2 ~ 30 seconds) read the absorbance value, at least read 4 points, to get 3 △ A; generally will be read many times in a row for the least-squares processing, readings are too short for the interval for the rate of time (TR) multi-point method of processing, are taken as part of the readings of the linear reaction, to find out the rate of the reaction per unit of time △ A / MIN. /This method must be based on the zero-level reaction, because only in the zero-level reaction, the absorbance change per unit time (reaction rate ΔA/MIN) is proportional to the enzyme activity. This method relatively reduces the analytical error and greatly improves the speed and accuracy of analysis. However, semi-automatic biochemical analyzers use continuous monitoring of a single sample, which is quite time-consuming. To apply the continuous monitoring method, first of all, we should prepare samples with high, medium and low concentrations within the linear range, and then make the reaction time curve to understand the whole process of the reaction under different concentrations, and determine the delay time and linear monitoring period.
2. Two-point rate method is the so-called proposed one-level rate method. In the reaction to select two time points T 1, T 2, read absorbance A 1, A2, calculate (A2-A1), (T2-T1) = △ A, △ T. This method and the end point of the two-point method of the difference between the two points: the latter reading point of the reaction did not reach the end point, the rate of the calculation of the results. It is compared with the continuous monitoring method, the disadvantage lies in the artificial determination of T 1, T 2, more uncertainty, can not guarantee that the reaction is linear during T 1-T 2, affecting the accuracy of the results; should be done before the routine determination of the pre-test to determine the linear time period. If the reaction is not linear within the selected time period (e.g., the zero-level reaction period is short and the instrument cannot be set or measured), the end-point method can only be used instead. The advantage is that the method is simple; lower enzyme activity, the determination of the absorbance value is small, can increase the measurement time period without the limitations of the instrument to continuously monitor the time point, reduce the reading error.
3. The Rate B method performs two reaction-related rate measurements in one channel at a time. It can be both the two test measurements, can also be used for interference or / and sample blanks automatically compensated for the latter use of the principle is: the use of the instrument's microcomputer automated processing, in the first reaction (interference reaction) has been to maintain the linearity of the premise, can be deducted from the second reaction (the main reaction) rate of the rate of the first reaction rate of the continuation of the impact. Such as for the elimination of bilirubin converted to biliverdin absorbance drop, the negative interference on the determination of creatinine picric acid method. Some instruments (e.g., HITACHI series) are set up for this method.
(C) blank (BLANK) correction
In the spectrophotometric method, often use the blank solution to adjust the absorbance of the instrument's zero point, or used to offset some of the interfering factors of the determination. In the determination of biochemical analyzers, in addition to the use of dual or multiple wavelengths, two-point method to eliminate background interference, often to use a special blank determination, in order to determine the absorbance from the sample to deduct its influence. The correct choice of blank calibration, to improve the accuracy plays an important role.
1. Reagent blanks are generally in the method type and calibration mode, that is, divided into two categories with or without reagent blanks. Reagent blanks are measured alone or in conjunction with calibration, and need to be pre-selected to load deionized water sample cups or reagent blank racks. The absorbance at each measurement point for calibration or patient samples is subtracted from the reagent blank absorbance or blank rate value at the corresponding measurement point. The method without reagent blank, more directly to the reaction cup of water blank as the determination of the reference value.
In many instruments, the reagent blank determination is similar to the calibration determination, not in the patient sample determination of real-time determination. Therefore, it is necessary to pay attention to the frequency of its determination to avoid calculation errors caused by changes in reagent blanks due to changes in reagent lot number or quality.
2. Sample blank Mainly to eliminate the sample itself turbidity or chromaticity of the interference. Often use the blank channel method, determination of the corrected results = color reaction channel results - blank channel results. Most of the instruments must be occupied in addition to the measurement channel, the analysis speed is halved, but the accuracy of the interference should be higher than the two-point endpoints
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