WO2018171619A1 - 一种硫氧还蛋白还原酶活性检测方法、检测设备及其操作方法 - Google Patents

一种硫氧还蛋白还原酶活性检测方法、检测设备及其操作方法 Download PDF

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WO2018171619A1
WO2018171619A1 PCT/CN2018/079787 CN2018079787W WO2018171619A1 WO 2018171619 A1 WO2018171619 A1 WO 2018171619A1 CN 2018079787 W CN2018079787 W CN 2018079787W WO 2018171619 A1 WO2018171619 A1 WO 2018171619A1
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reagent
target
reaction
sampling
reaction cup
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PCT/CN2018/079787
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English (en)
French (fr)
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尹汉维
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南京凯熙医学科技有限公司
武汉尚宜康健科技有限公司
凯熙医药（武汉）股份有限公司
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Priority claimed from CN201710170517.9A external-priority patent/CN108627659A/zh
Priority claimed from CN201710172659.9A external-priority patent/CN108627469B/zh
Application filed by 南京凯熙医学科技有限公司, 武汉尚宜康健科技有限公司, 凯熙医药（武汉）股份有限公司 filed Critical 南京凯熙医学科技有限公司
Priority to US16/496,207 priority Critical patent/US20200190556A1/en
Publication of WO2018171619A1 publication Critical patent/WO2018171619A1/zh

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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G01N35/02—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor using a plurality of sample containers moved by a conveyor system past one or more treatment or analysis stations
    • G01N35/04—Details of the conveyor system
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/26—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving oxidoreductase
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G01N35/00584—Control arrangements for automatic analysers
    • G01N35/00722—Communications; Identification
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G01N35/10—Devices for transferring samples or any liquids to, in, or from, the analysis apparatus, e.g. suction devices, injection devices
    • G01N35/1004—Cleaning sample transfer devices
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G01N35/10—Devices for transferring samples or any liquids to, in, or from, the analysis apparatus, e.g. suction devices, injection devices
    • G01N35/1009—Characterised by arrangements for controlling the aspiration or dispense of liquids
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G01N2035/00465—Separating and mixing arrangements
    • G01N2035/00514—Stationary mixing elements
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G01N35/02—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor using a plurality of sample containers moved by a conveyor system past one or more treatment or analysis stations
    • G01N35/04—Details of the conveyor system
    • G01N2035/0401—Sample carriers, cuvettes or reaction vessels
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G01N35/02—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor using a plurality of sample containers moved by a conveyor system past one or more treatment or analysis stations
    • G01N35/04—Details of the conveyor system
    • G01N2035/0401—Sample carriers, cuvettes or reaction vessels
    • G01N2035/0437—Cleaning cuvettes or reaction vessels
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G01N35/02—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor using a plurality of sample containers moved by a conveyor system past one or more treatment or analysis stations
    • G01N35/04—Details of the conveyor system
    • G01N2035/0439—Rotary sample carriers, i.e. carousels
    • G01N2035/0441—Rotary sample carriers, i.e. carousels for samples
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G01N35/02—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor using a plurality of sample containers moved by a conveyor system past one or more treatment or analysis stations
    • G01N35/04—Details of the conveyor system
    • G01N2035/0439—Rotary sample carriers, i.e. carousels
    • G01N2035/0446—Combinations of the above
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
    • G01N2333/90—Enzymes; Proenzymes
    • G01N2333/902—Oxidoreductases (1.)
    • G01N2333/90212—Oxidoreductases (1.) acting on a sulfur group of donors (1.8)

Definitions

  • the invention belongs to the field of enzyme activity detection, and particularly relates to a biochemical detection method, a detection device and an operation method thereof for the activity of thioredoxin reductase (TR) in human blood.
  • TR thioredoxin reductase
  • TR thioredoxin reductase
  • TR detection equipment can be applied to early screening of tumors in the physical examination population, tumor efficacy monitoring, recurrence warning and health management in hospitalized population, which has great market demand and development potential.
  • Patent ZL201080049877.X Method and kit for determining the activity of thioredoxin reductase in a sample and application
  • manual step-by-step operation such as: manual operation of a microplate reader, manual sample loading, Hand-operated, protected from light, manually placed on the shaker to achieve shake, manual sample loading, etc.; the various conditions of the sample are also designed for manual operation process and dosage.
  • Manual manual operation includes large manual error; various operations are non-coherent, and intermediate errors are easy to occur; long operation time is not conducive to various problems such as large-scale clinical detection applications.
  • TR detection kit in the prior art is a preliminary research and development product, and its detection steps, actions, fluxes and the like have certain limitations, so there is still room for improvement and correction in terms of detection speed and detection accuracy.
  • TR thioredoxin reductase
  • TR thioredoxin reductase
  • the object of the present invention is to provide a new fully automated thioredoxin reductase activity detecting method, detecting device and operating method thereof, thereby saving detection time and detecting steps.
  • the invention solves the problem of manually detecting thioredoxin reductase in human blood in the prior art, and can improve the detection efficiency and cost saving of thioredoxin reductase detection in clinical.
  • the invention realizes the full automatic operation of the "TR activity detection" on the biochemical detection device for the first time; the data directly obtained by the invention meets the requirements of the national detection standard, and can ensure that the detection data result reflects the early warning function of the detection technology (conformity Distribution of national data indicators).
  • a method for detecting thioredoxin reductase activity comprising: dosing, disposing a working solution, an inhibitor solution, and a mixed reagent; adding a sample, adding 50 uL to 70 uL of the working solution to the control reaction cup Add 50uL-70uL inhibitor solution to the reaction cup of the experimental group; add 10uL-30uL sample to the reaction cup of the control group and the reaction cup of the experimental group respectively; the amount of the sample is the volume required for the optimal work of the automated collaborative testing equipment. design.
  • the sample is incubated, and the control reaction cup and the experimental reaction cuvette are placed at a temperature of 30 ° C to 40 ° C for the first predetermined time in a dark environment.
  • the step of disposing the working fluid comprises: taking hydroxymethylaminomethane hydrochloride, morpholinyl propanesulfonic acid, disodium hydrogen phosphate citrate buffer system and disodium hydrogen phosphate according to a ratio of 1:1:2:4. Potassium dihydrogen phosphate buffer solution; mixing of automated synergistic detection equipment: trishydroxymethylaminomethane hydrochloride, morpholinyl propanesulfonic acid, disodium hydrogen phosphate citrate buffer system and disodium hydrogen phosphate disodium hydrogen phosphate buffer solution Evenly.
  • the automated synergistic detection device tris (hydroxymethyl) aminomethane hydrochloride has a pH of 5.5-7.2 and a concentration of 0.025-0.125 mol/L; the automated synergistic detection device has a concentration of morpholylpropanesulfonic acid of 0.25 mol/L.
  • the automated synergistic detection equipment pH of dihydrogen phosphate citrate buffer system is 2.2-8.0, the concentration is 0.2mol / L; the automated synergistic detection equipment pH of the potassium phosphate dihydrogen phosphate buffer solution is 4.9-8.2, The concentration is 1-15 mol/L.
  • the step of configuring the inhibitor solution by the automated collaborative detection device comprises: mixing the automated synergistic detection device working fluid and the inhibitor in a ratio of 1:1 to 1:5 to form an automated synergistic detection device inhibitor solution;
  • the synergistic detection device inhibitor solution is uniformly mixed, and the automated synergistic detection device inhibitor is a thioredoxin reductase inhibitor compound.
  • the step of configuring the mixing reagent by the automated collaborative testing device comprises: mixing the automated collaborative testing device reagent A and the automated collaborative testing device reagent B in a ratio of 1:4-1:8 to form an automated synergistic detecting device mixing reagent; Mix the automated synergistic detection equipment mixing reagents uniformly; the automated synergistic detection equipment A reagent is 5,5-dithiobis(2-nitrobenzoic acid) or substituted 6,6'-dinitro-3,3' - Dithiobenzoic acid; automated synergistic detection device B reagent is nicotinamide adenine dinucleotide phosphate.
  • the automated collaborative detection device has a predetermined temperature of 30-40 °C.
  • the automated collaborative detection device has a first predetermined time of 8-20 minutes.
  • the automated collaborative detection device has a first predetermined time of 10 minutes.
  • the automated collaborative detection device has a predetermined wavelength of 405 nm to 450 nm.
  • the automated collaborative detection device has a second predetermined time of 20-30 cycles.
  • the invention provides a method for detecting human peripheral blood thioredoxin reductase activity, which automatically samples and mixes reagent A and reagent B into a mixed reagent on an automated collaborative detection device, and automatically joins the sample through a synergistic detecting device.
  • the mixing and stirring operation replaces the work of separately adding the reagent A and the reagent B for manual repeated mixing and stirring, thereby improving the working efficiency.
  • the detection method can meet the requirements of the method for selecting the working fluid selection by setting the sample/reagent volume of the synergistic detecting device suitable for the thioredoxin reductase activity described above, and the method of the present invention is described above.
  • the method When used in a synergistic detecting device for thioredoxin reductase activity, the method includes a sample loading operation method, a light shielding operation request method, and an operation request method for reagent mixing, and the like, and an intelligent instruction method for a drive system.
  • the intelligent instructions such as the number of cycles in each cycle group and the time of each cycle in the operation of the collaborative detection device, and the requirements for the operation process are all detection methods that are connected with the human peripheral blood TR function detection method.
  • the method of the present invention is a method for realizing the requirement of TR enzymatic detection function by using a synergistic detecting device for thioredoxin reductase activity described above, and is used for the synergistic detection of a thioredoxin reductase activity described above.
  • a biochemical detection device for thioredoxin reductase detection comprising:
  • the accommodating device is configured to hold a plurality of reagents and/or samples, and under the driving action of the driving device, periodically rotate around the axis to rotate the target reagent and/or the target sample to the target filling hole;
  • reaction device for holding a plurality of cuvettes, and under the driving action of the driving device, periodically rotating around the axis to rotate the target cuvette to the target sampling hole;
  • a sampling device configured to periodically rotate around the axis under the driving of the driving device, and add the target reagent and/or the target sample collected from the target sampling hole to the target reaction cup corresponding to the target filling hole ;
  • a state sensing device for detecting reaction rotation state information of the reaction device, accommodation rotation state information of the accommodation device, and reagent state information of the sampling device;
  • the main control system is respectively connected to the state sensing device and the driving device, and is configured to generate and send a corresponding control instruction to the driving device based on the reaction rotation state information, the accommodation rotation state information, and the reagent state information;
  • the driving device is respectively connected to the sampling device, the accommodating device and the reaction device, and is configured to control the sampling device, the reaction device and the accommodating device to perform corresponding operations based on the received control instruction.
  • a method of operating a biochemical detection device comprising:
  • the state sensing device detects that the target reaction cup in the reaction device rotates to the target liquid adding hole and the reagent state information of the sampling device is the reagent collecting completion state, generating a reagent adding instruction
  • the driving device controls the sampling device to rotate to the target liquid adding hole of the reaction device based on the received reagent adding instruction, and adds the target reagent and/or the target sample to the target reaction cup corresponding to the target liquid adding hole;
  • the driving device controls the sampling device to rotate to the target sampling hole of the loading device to collect the target reagent and/or the target sample based on the received reagent collection instruction.
  • the present invention proposes a fully automatic biochemical detection device for detecting thioredoxin reductase (TR) in human blood, which realizes full automation of biochemical detection equipment, improves detection efficiency, and saves cost. .
  • FIG. 1 is a schematic flow chart of a method for detecting thioredoxin reductase activity of the present invention
  • FIG. 2 is a schematic structural view of a corresponding detection device for detecting a thioredoxin reductase activity of the present invention
  • FIG. 3 is a schematic flow chart of a method for detecting thioredoxin reductase activity according to a specific embodiment of the present invention
  • FIG. 4 is a schematic flow chart showing another method for detecting thioredoxin reductase activity according to a specific embodiment of the present invention
  • FIG. 5 is a flow chart of step S1 in the flow chart shown in Figure 4;
  • Figure 6 is a flow chart of step S2 in the flow chart shown in Figure 4;
  • FIG 7 is a flow chart of step S3 in the flow chart shown in Figure 4;
  • Figure 8 is a schematic diagram showing the system architecture of the biochemical detection device for thioredoxin reductase (TR) detection of the present invention
  • Figure 9 is a schematic view showing the mechanical structure of a biochemical detecting device for thioredoxin reductase (TR) detection of the present invention.
  • FIG. 10 is a schematic flow chart showing the operation method of the biochemical detecting device for detecting thioredoxin reductase (TR) according to the tenth embodiment of the present invention
  • Figure 11 is a flow chart showing the operation method of the biochemical detecting device for detecting thioredoxin reductase (TR) according to the eleventh embodiment of the present invention
  • Figure 12 is a flow chart showing the operation method of the biochemical detection device for thioredoxin reductase (TR) detection according to the twelfth embodiment of the present invention
  • Figure 13 is a flow chart showing the operation method of the biochemical detection device for thioredoxin reductase (TR) detection according to the thirteenth embodiment of the present invention
  • FIG. 14 is a schematic diagram of the detection principle of a detecting device according to an embodiment of the present invention.
  • the detection method used in the "TR activity detection kit" in the prior art (hereinafter referred to as the initial generation detection method) has three technical limitations, and thus cannot be applied to an automated collaborative detection device:
  • the initial detection method includes 7 steps and actions, which is not difficult for the manual detection of the test personnel, but is too cumbersome for the automated collaborative detection device, so the operation time of the device is significantly increased, and the detection efficiency is lowered;
  • the initial detection method requires two inspectors to cooperate with each other during the operation, and the automated collaborative detection equipment is designed to allow a single inspector to operate the instrument completely, so the requirements of the two do not match.
  • the reagent A in the present invention is an agent for detecting the activity of thioredoxin reductase
  • the reagent B is an agent for detecting the activity of thioredoxin reductase.
  • the working fluid is usually a buffer, and is mainly used as a buffer for detecting thioredoxin reductase activity.
  • the reagent A and the reagent B in the invention have all passed the expert certification and examination by the State Food and Drug Administration, and obtained the medical device registration certificate (the certificate number is the food and medicine supervision (standard) word 2013 No. 2401815 and the country Reagent A and reagent B in the thioredoxin reductase (TR) activity detection kit of Food and Drug Administration (Jun) 2014 3400264), the working fluid in the present invention is the above-mentioned "thioredoxin reduction"
  • the reagent D in the enzyme (TR) activity detecting kit, and the inhibitor in the present invention is the reagent C in the above-mentioned "thioredoxin reductase (TR) activity detecting kit".
  • Thioredoxin reductase is a reduced coenzyme II (NADPH)-dependent dimeric selenoenzyme containing a flavin adenine dinucleotide (FAD) domain, which is reduced with thioredoxin.
  • the sex coenzyme II together constitutes a thioredoxin system.
  • Thioredoxin reductase is overexpressed in cells with abnormally active proliferation, and this enzyme has physiological functions such as priming cell abnormal proliferation, initiation of apoptosis inhibition system and the like to form a tumor.
  • TR activity is highly correlated with the degree of abnormal hyperplasia of the tumor. Therefore, the detection of thioredoxin reductase has an important effect on tumors.
  • Fig. 1 is a schematic flow chart showing the method for detecting thioredoxin reductase activity of the present invention.
  • the method for detecting thioredoxin reductase activity comprises:
  • Step S1 dosing, disposing a working solution, an inhibitor solution, and a mixing reagent.
  • the working fluid is usually a buffer, and the working fluid is not particularly limited in its concentration, and is preferably formulated to have a 1X working fluid concentration.
  • the working fluid configuration process is: according to the ratio of 1:1:2:4, the reagent TrisHCL (0.025-0.125 mol / L, pH 5.5-7.2), morpholinyl propanesulfonic acid (0.25mol/L), hydrogen phosphate disodium citrate buffer system 0.2mol/L, and hydrogen phosphate disodium phosphate potassium dihydrogen phosphate buffer solution 1-15mol/L; wherein, the pH of the disodium hydrogen phosphate citrate buffer system is 2.2-8.0; pH of diammonium phosphate dipotassium phosphate buffer solution is 4.9-8.2; then buffer TrisHCL (trishydroxymethylaminomethane hydrochloride), morpholinyl propanesulfonic acid, disodium hydrogen phosphate citrate The system was uniformly mixed with a potassium hydrogen phosphate disodium hydrogen phosphate buffer solution.
  • the working solution and the inhibitor are mixed in a ratio of 1:1 to 1:5 to form an inhibitor solution; the inhibitor solution is uniformly mixed; wherein the inhibitor is a thioredoxin reductase inhibitor compound. It can be a chemical monomer selenium compound.
  • the ratio of the working fluid and the inhibitor is preferably a ratio of 1:3, and the working fluid and the inhibitor mixed at the ratio are the most economical ratio combination, that is, the subsequent human peripheral blood thioresin reductase
  • the activity detection is more accurate and is the most economical combination of reagents.
  • the reagent A and the reagent B are mixed in a ratio of 1:2-1:8 to form a mixed reagent; specifically, the mixing ratio of the reagent A and the reagent B is in the range of 1:2-1:8, preferably 1:4-5.
  • the ratio of the reagent A and the reagent B are mixed, and the mixed reagent formed by the mixing at this time has higher accuracy in the detection, wherein the A reagent is 5,5-dithiobis(2-nitrobenzoic acid) or substituted 6 , 6'-dinitro-3,3'-dithiobenzoic acid; B reagent is nicotinamide adenine dinucleotide phosphate.
  • the mixed working solution evenly; mix the inhibitor solution evenly; mix the mixing reagent evenly.
  • the mixed working solution, the inhibitor solution, and the mixed reagent are separately placed, and usually placed in a reagent tank to facilitate subsequent detection.
  • Step S2 adding a sample, adding 50 uL-70 uL of the working solution to the control reaction cup; adding 50 uL-70 uL of the inhibitor solution to the reaction cup of the experimental group; adding 10 uL to 30 uL of the sample to the reaction cup of the control group and the reaction cup of the experimental group;
  • the reaction cups of the control group and the experimental group were set at intervals, for example, the odd numbered reaction cups were the control reaction cups, and the even numbered reaction cups were the experimental group reaction cups.
  • the amount of the working fluid added to the reaction cup of the control group and the reaction cup of the experimental group is equivalent.
  • 50 uL-70 uL of the working solution is generally added to the reaction cup of the control group and the reaction cup of the experimental group.
  • 50-60 uL of working fluid is added to the control cup and the experimental set of reaction cups.
  • an equal amount of working fluid is added to the reaction cup of the control group and the reaction cup of the experimental group, so that the test data of the control group and the experimental group are comparable, and the data after the detection can be calculated.
  • the sample loading time of each sample is about 10 minutes, that is, 27 cycles, 22.5 seconds per cycle. After the sample is added, the sample of this group can be incubated, and then the other sample can be further loaded.
  • Step S3 incubating, in a dark environment, the control reaction cup and the experimental reaction cup are placed at a temperature of 30 ° C - 40 ° C for a first predetermined time; wherein, the predetermined temperature is 30 ° C - 40 ° C; The predetermined time is 8-20 minutes, preferably 10 minutes.
  • control reaction cup and the experimental reaction cup are placed at a temperature of 30 ° C to 40 ° C for 8-20 minutes in the dark, preferably 10 minutes, because the detection method can be combined with the detection.
  • the device can be used for automated detection on a synergistic detection device for thioredoxin reductase activity at a predetermined time of 10 minutes. This group of samples can be assayed after a set of samples has been incubated.
  • step S4 it is determined that 100 uL-150 uL of the mixed reagent is added to the reaction cup of the control group and the reaction cup of the experimental group, respectively; and the absorbance value in the second predetermined time period is measured at a predetermined wavelength.
  • the absorbance value in the second predetermined time period is measured at a predetermined wavelength.
  • 120uL can be added in the reaction cup of the control group and the reaction cup of the experimental group.
  • the mixed reagent continuously measures the absorbance value of the sample at a wavelength of 405 nm to 450 nm for 7.5-11.25 minutes, that is, 20-30 cycles.
  • the mixed reagent is directly added to the reagent to be measured, thereby achieving mixed mixing, which is higher than that in the prior art.
  • Reagent A and reagent B were separately stirred, and then separately applied, a part of the steps were omitted, the detection time was saved, and automatic detection on the synergistic detection device of thioredoxin reductase activity was realized, and the detection was improved. effectiveness.
  • sample in the present invention refers to any tissue derived from a living organism or a portion separated therefrom, and the sample is preferably blood, body fluid, tissue homogenate, and most preferably blood, wherein the blood may be a component such as serum, plasma or the like.
  • the detection method of the invention can be applied to "a thioredoxin activity detecting device", as shown in FIG. 2, the structure of the device is specifically:
  • Testing equipment includes:
  • the apparatus 2 includes a sample reagent disk 2-1 and a sample tube fixing member 2-2, a working fluid/inhibitor solution fixing member 2-3, and a mixed reagent fixing member 2 which are uniformly distributed in the circumferential direction of the sample reagent disk 2-1, respectively.
  • the sample tube fixing member 2-2 may be a ring-shaped structural member having a plurality of holes arranged around the circumference of the sample reagent disk, and placed in the sample reagent tray to place the sample tube, for example, a test tube rack, a sample tube
  • the number of holes in the fixing member is preferably 40;
  • the structure of the working fluid/inhibitor solution fixing member 2-3 and the mixed reagent fixing member 2-4 is similar to that of the sample tube fixing structure for respectively placing the working fluid/inhibitor solution bottle Or mixing the reagent bottles, the number is preferably 30 or 40, and each of the sample tube fixing members 2-2, each of the working fluid/inhibitor solution fixing members 2-3 and each of the mixing reagent fixing members 2-4 is Center of sample reagent plate Center, and sequentially distributed from inside to outside along the radius of the sample reagent disk.
  • the reaction device 3 is periodically rotated around the axis under the driving action of the driving system; the reaction device 3 includes: a reaction disk 3-1 and a plurality of cuvette fixing members 3-2 uniformly distributed along the circumferential direction of the reaction disk, and the reaction
  • the cup fixing part 3-2 may be an annular structural part having a plurality of holes arranged around the circumferential direction of the reaction disk, and placed in the reaction tray to accommodate or place the cuvette, for example, a test tube rack, and the number of the reaction cups is 81, can be divided into 9 groups.
  • a sampling device 4 for collecting reagents and/or samples from the accommodating device 2 into the reaction device 3 based on the periodic rotation of the accommodating device 2 and the reaction device 3;
  • the sampling device 4 is optionally a sampling needle;
  • the drive system is connected to the accommodating device 2, the reaction device 3 and the sampling device 4 for controlling the accommodating device 2, the reaction device 3 and the sampling device 4 to perform corresponding operations.
  • the hydraulic device 5 includes a vacuum pump 5-1, a vacuum tank 5-2, and a liquid path mechanism 5-3.
  • the liquid path mechanism includes a pipe and a valve disposed on the pipe, and the liquid path mechanism is also connected to the sampling device and the automatic cleaning device.
  • a vacuum pump 5-1 for adjusting the air pressure in the vacuum tank 5-2 so that the air pressure in the vacuum tank 5-2 reaches a preset air pressure
  • the vacuum tank 5-2 is configured to control the operation of the liquid path mechanism at a preset air pressure to perform an ascending, descending, and rotating operation on the sampling device, and to cause the liquid to enter or exit the automatic cleaning device.
  • the automatic cleaning device 6 is connected to the hydraulic device 5 for cleaning the reaction cup in the reaction device 3 based on the control of the hydraulic device 5; the automatic cleaning device 6 may be a cleaning needle capable of sucking liquid into the accommodation space inside the cleaning needle And then released into the reaction cup to clean the reaction cup, and then the washed waste liquid is sucked out from the reaction cup and discharged to a waste water tank (not shown), and the waste water pool can be disposed outside the detection device.
  • the stirring device 7 is connected to the driving system for agitating the mixed solution formed by the reagent and the sample, and uniformly stirring after completion of each loading step.
  • the agitation means may be a stirring needle.
  • the temperature control device 8 is located below the plurality of cuvette fixing members for controlling the temperature of the cuvette in the cuvette holding member to be maintained at the set experimental temperature.
  • the temperature control device 8 is a temperature control tank having a trough-like structure such that 81 cuvettes can be located inside the trough structure to keep the ambient temperature inside the reaction device at the reaction temperature and the incubation temperature during the reaction.
  • the optoelectronic device 9 is disposed on the upper surface of the casing 1 for controlling the optical path and the wavelength, providing illumination for the reaction process of the sample and the reagent, and continuously measuring the absorbance value of the sample.
  • the photovoltaic device is a photovoltaic box.
  • the syringe 10 is disposed on the upper surface of the housing 1.
  • the sample reagent tray is provided with a sample reagent disk cover to provide an experimental environment for the sample disk to be closed for TR activity detection;
  • the sample reagent disk cover is provided with a sample sampling hole, a working fluid/inhibitor solution sampling hole and a mixed reagent.
  • a sampling hole a sampling hole; a first liquid adding hole and a second liquid adding hole are arranged on the reaction disk; the sample sampling hole, the working liquid/inhibitor solution sampling hole, the mixed reagent sampling hole, the first liquid adding hole and the second liquid adding hole are both Located on the same circle centered on the sampling device, the sampling device is along the circumference of the circle in the sample sampling hole, the working fluid/inhibitor solution sampling hole, the mixed reagent sampling hole, the first dosing hole and the second dosing hole. Do periodic exercises.
  • the interval between the first liquid addition hole and the second liquid addition hole may be set to be less than the distance of the number of reaction cups of each set of reaction cups, preferably, the first liquid addition hole and the second liquid addition hole The interval between them can be set to a distance of 7 to 11 cuvettes, and more preferably, a distance set to 8 cuvettes.
  • the embodiment of the invention further includes a host computer, and the upper computer is connected with the hydraulic device and the driving system for transmitting an operation instruction to the hydraulic device and the driving system, so that the driving system controls the receiving device 2 and the reaction device 3 to perform periodic rotation when needed During sampling, the drive system receives the command from the upper computer to control the rotation of the accommodating device.
  • a sample tube fixing member, a working fluid/inhibitor solution bottle fixing member and a mixed reagent bottle fixing member are rotated to respectively correspond to
  • the sampling device will be lowered to the sample tube fixing member and the working fluid/inhibitor solution under the control of the hydraulic device.
  • the sample, the working liquid/inhibitor solution or the mixed reagent is collected from the sample tube, the working fluid/inhibitor solution bottle or the mixed reagent bottle, and then rises under the control of the hydraulic device, and then Move to the position of the first dosing hole or the second dosing hole and drop under the action of the hydraulic device , the collected sample, working fluid/inhibitor solution or mixed reagent is placed in the reaction cup at the first filling hole or the second filling hole position, and at the same time, the reaction device rotates one space to make the first filling liquid
  • the hole and the second liquid addition hole respectively correspond to the next reaction cup that needs to be added, and then continue to complete the collection of the next cycle and dosing.
  • a receiving device for accommodating the sample and the reagent, and periodically rotating around the axis under the driving action of the driving system
  • the reaction device for accommodating the reaction cup and the experimental cup, and the driving action of the reaction cup and the experimental cup in the driving system Next, periodically rotating around the axis; sampling means for collecting reagents and/or samples from the receiving device into the reaction device based on the periodic rotation of the receiving device and the reaction device; the driving system, respectively, and the receiving device, the reaction
  • the device is coupled to the sampling device for controlling the operation of the receiving device, the reaction device, and the sampling device.
  • the following methods are applicable to both the experimental group and the control group, and the detection methods include:
  • 3 and 4 are schematic flow charts showing the method for detecting thioredoxin reductase activity according to an embodiment of the present invention.
  • a method for detecting thioredoxin reductase activity comprises:
  • the driving system controls the sampling device to sequentially collect the working fluid/inhibitor solution from the sample reagent tray, and controls the sampling device to sequentially add the collected working fluid/inhibitor solution to the first group of reaction cups until the first One cycle group ends;
  • step S1 includes:
  • the driving system controls the reaction disk to rotate, so that the first reaction cup in the first group of reaction cups is located at the first liquid filling hole position;
  • the driving system controls the sampling device to rotate to the position of the sampling hole of the working fluid/inhibitor solution, and collects the working fluid/inhibitor solution from a working fluid/inhibitor solution bottle of the sample reagent tray;
  • the driving system controls the sampling device to rotate to the position of the first filling hole, and the collected working fluid/inhibitor solution is added into the reaction cup at the position of the first filling hole;
  • the driving system controls the sampling device and the reaction tray to repeat the adding step of the working fluid/inhibitor solution until each of the first reaction cups is added with the working fluid/inhibitor solution, and the first cycle group ends.
  • each of the preset angles may be set as the interval between the two cuvette fixing members, such that the setting can be such that each of the cuvettes in the first set of cuvettes is sequentially located in the first plus for each predetermined angle.
  • the drive system controls the sampling device to sequentially collect the working fluid/inhibitor solution from the sample reagent disk, and controls the sampling device to collect the working fluid/inhibition.
  • the solution is sequentially added to the second set of reaction cups;
  • the drive system controls the sampling device to sequentially collect samples from the sample reagent disk, and sequentially controls the sampling device to add the collected samples to the first set of reaction cups until the first cycle.
  • the second cycle group ends;
  • step S2 includes:
  • the driving system controls the reaction disk to rotate, so that the first reaction cup in the second group of reaction cups is located at the first liquid filling hole position;
  • the driving system controls the sampling device to rotate to the sampling hole position of the working fluid/inhibitor solution, collects the working fluid/inhibitor solution from a working fluid/inhibitor solution bottle of the sample reagent tray, and controls the sampling device to rotate to the first At a liquid filling position, the collected working fluid/inhibitor solution is added to the reaction cup at the first filling hole position;
  • the driving system controls the reaction disk to rotate, so that the first reaction cup in the first group of reaction cups is located at the second liquid filling hole position;
  • the driving system controls the sampling device to rotate to the position of the sample sampling hole, collects the sample from a sample tube of the sample reagent disk, and controls the sampling device to rotate to the position of the second liquid filling hole, and adds the collected sample to the second plus Inside the reaction cup at the location of the liquid well;
  • the reaction disk may or may not rotate. If the third cycle of the second cycle group is selected not to rotate, the rotation may be selected in the next cycle, but Considering the accuracy and convenience of the program control, the reaction disk preferably does not rotate; and if the sample reagent disk needs to be rotated during this cycle, it is also possible to select to rotate in this cycle or in the next cycle.
  • the drive system controls the sampling device to sequentially collect the working fluid/inhibitor solution from the sample reagent disk every two cycles, and controls the sampling device to collect the working fluid/inhibitor The solution is sequentially added to the third set of reaction cups;
  • the drive system controls the sampling device to sequentially collect samples from the sample reagent disk every two cycles, and controls the sampling device to sequentially add the collected samples to the second set of reaction cups;
  • the drive system controls the sampling device to sequentially collect the mixed reagent from the sample reagent disk every two cycles, and controls the sampling device to sequentially add the collected mixed reagents to the first set of reaction cups until The third cycle group ends.
  • the first set of reaction cups is filled with liquid and begins to enter the incubation time.
  • step S3 includes:
  • the driving system controls the reaction disk to rotate, so that the first reaction cup in the third group of reaction cups is located at the first liquid addition hole or the second liquid addition hole position;
  • the driving system controls the sampling device to rotate to the sampling hole position of the working fluid/inhibitor solution, collects the working fluid/inhibitor solution from a working fluid/inhibitor solution bottle of the sample reagent tray, and controls the sampling device to rotate to the first Adding the working liquid/inhibitor solution to the reaction cup at the first liquid addition hole or the second liquid addition hole position at a liquid addition hole or a second liquid addition hole position;
  • the driving system controls the reaction disk to rotate, so that the first reaction cup in the second group of reaction cups is located at the first liquid addition hole or the second liquid addition hole position;
  • the driving system controls the sampling device to rotate to the position of the sample sampling hole, collects the sample from a sample tube of the sample reagent disk, and controls the sampling device to rotate to the position of the first liquid filling hole or the second liquid filling hole, and the collecting is performed. a sample is added to the cuvette at the first or second dosing position;
  • the driving system controls the reaction disk to rotate, so that the first reaction cup in the first group of reaction cups is located at the first liquid addition hole or the second liquid addition hole position;
  • the driving system controls the sampling device to rotate to the position of the mixed reagent sampling hole, collects the mixed reagent from a mixed reagent bottle of the sample reagent tray, and controls the sampling device to rotate to the position of the second filling hole, and the collected mixed reagent Adding into the reaction cup at the first filling hole or the second filling hole position;
  • the working fluid/inhibitor solution, the sample and the mixed reagent are respectively added to each reaction cup in turn, until the whole plate detection is completed or the detection is stopped;
  • the dosing is cycled according to the three-cycle dosing step of the third cycle group until the working fluid/inhibitor solution, the sample and the mixed reagent are added to all the cuvettes or the detection is stopped.
  • step S4 specifically, starting from the first cycle of the fourth cycle group, the working fluid/inhibitor solution is sequentially collected every two cycles, and sequentially added to the fourth group of reaction cups; At the beginning of the second cycle, samples are taken sequentially, and the third set of reaction cups are sequentially added; from the third cycle of the fourth cycle group, the mixed reagents are sequentially collected every two cycles, and sequentially Add a second set of reaction cups;
  • the reaction device As long as the reaction device is in operation, the above step S3 can be repeated. It should be noted that after each working group is added with the working fluid/inhibitor solution, the sample and the mixed reagent, the reaction starts.
  • the reaction time is up to 22 cycles (22.5 s per cycle), preferably 20 cycles.
  • the hydraulic device can be sent by the upper computer to control the automatic cleaning device to remove the waste liquid after the reaction in the reaction cup, and the reaction cup is cleaned. After the cleaning is completed, the reaction cup is cleaned.
  • the operation of adding the working fluid/inhibitor solution, the sample, and the mixing reagent can be continued.
  • the mixture of the working fluid/inhibitor solution, the sample and the mixed reagent in the first set of reaction cups begins to react, at this time, in the next During the cycle, if the first set of reaction cups of the reaction disk are rotated to the dosing position, the sampling device is left in a waiting state.
  • the collaborative testing device performs a corresponding testing process
  • 81 reaction cups are divided into 9 groups, and each reaction has a total of 74 test cycles, including adding the first reagent to the 27 cycles of adding the sample, adding the sample to the joining 27 cycles of the two reagents, and 20 cycles of the reaction, each cycle takes 22.5 seconds, in the embodiment of the present invention
  • the sampling device takes the working fluid/inhibitor solution (working fluid or inhibitor solution) from the sampling hole position of the working fluid/inhibitor solution, and adds it to the No. 1 reaction cup of the reaction device, and then the reaction device rotates by 1 grid.
  • Spend 1 cycle (22.5s);
  • the working fluid/inhibitor solution is sequentially collected, and the working fluid/inhibitor solution is sequentially added to the 10-18 reaction cup;
  • the sampling device does not operate during the 12, 15, 18, 21, 24, 27, 30, 33, and 36 cycles.
  • the working fluid/inhibitor solution was collected sequentially at intervals of 3 cycles from the 37th cycle, and the reaction cups 19-27 were sequentially added; samples were sequentially collected every 3 cycles from the 38th cycle, and 10-19 were sequentially added.
  • the reaction cup was collected from the 39th cycle at intervals of 3 cycles, and the reaction cups 1-9 were sequentially added.
  • the working fluid/inhibitor solution is added to the No. 19 reaction cup; in the 38th cycle, the sample is added to the No. 10 reaction cup; in the 39th cycle, the mixed reagent is added to the No. 1 reaction cup; Add the working fluid/inhibitor solution to the No. 20 reaction cup; in the 41st cycle, add the sample to the No. 11 reaction cup; in the 42nd cycle, add the mixed reagent to the No. 2 reaction cup, and cycle accordingly, until the whole plate is completed or stopped. Detection
  • the cuvette group that has been previously reacted may enter the next round of inspection after washing.
  • the reaction disk can be set to rotate clockwise throughout the test
  • the reaction disk When the reaction disk is rotated clockwise, the reaction disk rotates clockwise in each cycle of the first cycle group, and the first reaction cup in the first group of reaction cups can be rotated clockwise or counterclockwise.
  • the reaction cup related to the liquid to be added cannot be rotated by a preset angle to each cycle to the first Or at the position of the second filling hole, it can be rotated clockwise to the position of the two filling holes, alternatively, it can be rotated counterclockwise.
  • the two disks work together.
  • the sample is added until the mixed reagent is added for 27 cycles (about 10min), in line with the medical requirements of TR activity detection; according to the above method to achieve detection of human blood samples, the background reduction of human blood samples by TR specific inhibitors is achieved, thereby ensuring that the detection method conforms to the detection results of TR enzyme activity and is related to the state Standard consistency (TR detection data for normal population is less than 4 units, and TR detection data for patients with high tumors is greater than 12 units).
  • the method for detecting thioredoxin reductase activity of the present invention can meet the requirements of automated detection by improving the primary detection method, and further has the following advantages:
  • the incubation time is significantly reduced, and the incubation time of a single sample is reduced from 30 minutes to 10 minutes, which can effectively reduce the detection time, so that every 40-50 samples
  • the detection can be completed within 1.5 hours, and the continuous detection of the synergistic detection equipment is realized, and the detection flux and speed requirements of the synergistic detection equipment for the thioredoxin reductase activity are met.
  • the detection action step is significantly reduced, from 7 action steps to 3-4 action steps, the detection step is optimized, and the instrument is reduced.
  • the action time and convenient instrument program operation so that a single tester can independently control the instrument to complete the entire test process.
  • the detection method of the present invention can be adapted to the action requirements, configuration requirements and instruction requirements of the TR-integrated detection device in conjunction with the thioredoxin reductase activity;
  • the clinical TR activity detection is performed on the synergistic detection device of thioredoxin reductase activity by the method of the invention, and is subjected to specific software (for details, please refer to another patent application: a thioredoxin reductase activity) Analytical method and system) treatment, can become the result of TR activity in clinical medicine, and meet the relevant requirements of the corresponding product "TR activity detection kit" and national medical device registration product standard: YZB / country (Q/CVH 001-2011 );
  • TR activity detection kit the certificate number is the food and medicine supervision (standard) word 2013 No. 2401815 and the national food medicine The supervision (probably) word 2014 3400264
  • TR detection results produced meet the relevant requirements of the corresponding products "TR activity detection kit” and national medical device registration product standards: YZB / country (Q / CVH 001-2011) .
  • the reagent A and the reagent B are automatically sampled and mixed into a mixed reagent by a synergistic detecting device for the thioredoxin reductase activity described above.
  • the synergistic detection device for thioredoxin reductase activity is automatically added to the sample for mixing and stirring, instead of adding reagent A and reagent B separately for manual mixing and stirring, thereby improving work efficiency.
  • Another object of the present invention is to provide a biochemical detection device for detecting the activity of thioredoxin reductase (TR) in human blood, which is used for realizing fully automatic biochemical detection, improving detection efficiency and saving cost.
  • TR thioredoxin reductase
  • Figure 8 is a schematic diagram showing the system architecture of the biochemical detection device for thioredoxin reductase (TR) detection of the present invention.
  • a first embodiment of the present invention provides a biochemical detection device, including: a receiving device 10, a reaction device 20, a sampling device 30, a state sensing device 50, a main control system 60, and a driving device 70.
  • the accommodating device 10 is configured to hold a plurality of reagents and/or samples, and under the driving action of the driving device 70, periodically rotate around the axis to rotate the target reagent and/or the target sample to the target liquid adding hole.
  • the reaction device 20 is configured to hold a plurality of cuvettes and, under the driving action of the driving device 70, periodically rotate around the axis to rotate the target cuvette to the target sampling hole.
  • the sampling device 30 is configured to periodically rotate around the axis under the driving of the driving device 70, and add the target reagent and/or the target sample collected from the target sampling hole to the target reaction corresponding to the target filling hole. In the cup.
  • the state sensing device 50 is configured to detect reaction rotation state information of the reaction device 20, accommodation rotation state information of the storage device 10, and reagent state information of the sampling device 30.
  • the main control system 60 is respectively connected to the state sensing device 50 and the driving device 70 for generating and transmitting corresponding control commands to the driving device 70 based on the reaction rotation state information, the accommodation rotation state information and the reagent state information;
  • Figure 9 is a schematic illustration of the mechanical structure of a biochemical detection device for thioredoxin reductase (TR) detection of the present invention.
  • the accommodating device 10 includes a accommodating tray 11 that is movably disposed to periodically rotate around the axis.
  • the accommodating tray 11 has a plurality of receiving fixtures 12 for holding reagent bottles and/or sample bottles.
  • At least one ring of the fixing member 12 is disposed on the accommodating disk 11 , and each of the rings has a plurality of accommodating fixing members 12 .
  • the number of accommodating fixtures 12 can be selected and set according to the specific needs of the user.
  • a plurality of accommodating fixing members 12 may be disposed uniformly along the edge of the accommodating tray 11, that is, each of the two accommodating fixing members 12 is spaced apart by a predetermined distance to form a uniform arrangement of at least one accommodating fixing member 12.
  • each receiving fixture 12 it can accommodate both the reagent bottle holder and the sample holder.
  • the reagent holder can be removed and replaced with the sample holder;
  • the sample holder can be removed and replaced with a reagent bottle holder, which realizes flexible exchange of the reagent position and the sample position, and can meet the different needs of different customers for the sample position and the reagent position.
  • the reaction apparatus 20 includes a reaction disk 21 which is movably disposed to periodically rotate around the axis.
  • a plurality of cuvette holders 22 for holding the cuvette.
  • At least one turn of the cuvette holder 22 is disposed on the reaction disk 21, and each of the rings has a plurality of cuvette holders 22.
  • the number of cuvettes 22 can be selected and set according to the specific needs of the user.
  • a plurality of cuvette holders 22 may be disposed uniformly along the edge of the reaction disk 21, that is, each of the two cuvette holders 22 is spaced apart by a predetermined distance to form a uniform arrangement of at least one cuvette holder 22.
  • the sampling device 30 includes a sampling rotating member 31, which is movably arranged for periodically rotating around the axis under the driving action of the driving device 70.
  • the sampling fixing member 32 is fixedly disposed on the sampling rotating member 31. Rotating under the belt action of the sampling rotary member 32; the sampling needle 33 has a fixed end at one end and a free end at the other end.
  • the main control system 60 generates a reagent addition command when receiving the reaction rotation state information when the target cuvette is rotated to the target liquid supply hole and the reagent state information is the reagent collection completion state; the driving device 70 receives the reagent addition instruction
  • the sampling device 30 is controlled to rotate to the target filling hole of the reaction device 20 and the target reagent and/or the target sample are added to the target reaction cup corresponding to the target filling hole.
  • the main control system 60 generates a reagent collection instruction when receiving the rotation state information when the target reagent and/or the target sample is rotated to the target sampling hole and the reagent state information is the reagent addition completion state; the driving device 70 is receiving After the reagent acquisition command, the sampling device 30 is controlled to rotate to the target sampling hole of the holding device 10 to collect the target reagent and/or the target sample.
  • the biochemical detection device for thioredoxin reductase (TR) detection provided by the third embodiment of the present invention further includes: a stirring device 40 for periodically rotating around the axis under the driving action of the driving device 70. The mixture formed in the target cuvette is stirred.
  • the agitating device 40 includes an agitating rotating member 41 fixedly disposed for periodically rotating around the axis under the driving action of the driving device 70.
  • the agitating fixing member 42 is fixedly disposed on the agitating rotating member. 31, the rotation of the agitating rotary member 42 is rotated; the agitating needle 43 has a fixed end at one end and a free end at the other end.
  • the main control system 60 generates a stirring control command when receiving the reagent state information as the reagent addition is completed;
  • the driving device 70 controls the stirring device 40 to rotate to the target liquid feeding hole to stir the mixed liquid in the target reaction cup.
  • the state sensing device 50 is further configured to detect the stirring state information of the stirring device 40; the main control system 60 And configured to generate a stirring reset command after receiving the stirring state information, and the driving device is connected to the stirring device 40 to control the stirring device 40 to reset.
  • the main control system 60 generates a reagent rotation command when receiving the reaction rotation state information when the number of times of rotating the predetermined angle is a predetermined number of times; after receiving the reagent rotation command, the accommodating device 10 controls the accommodating device 10 to rotate by a predetermined angle so that the next one The target cuvette is aligned with the target dosing hole.
  • the method further includes: a cleaning device 90, configured to input the liquid in the cleaning liquid into the target liquid adding hole The target cuvette is used to clean the target cuvette.
  • the cleaning device 90 includes: a hydraulic device 91 for controlling the liquid in the cleaning liquid to enter the target reaction cup or discharging the waste liquid in the target reaction cup to the target reaction cup; the cleaning assembly 92, and the hydraulic device 91 Connected, the target cuvette is cleaned by the hydraulic device 91.
  • the method further comprises:
  • the client 110 is connected to the main control system 60 for providing a user operation instruction input interface, and is configured to collect a cleaning instruction input by the user on the user operation instruction input interface, and send the cleaning instruction to the main control After receiving the cleaning command, the driving device 70 controls the cleaning device 90 to clean the target reaction cup corresponding to the target liquid filling hole.
  • the client 110 includes, but is not limited to, a host computer.
  • the method further comprises:
  • the temperature control device 80 under the driving action of the driving device 70, adjusts the experimental temperature of the reaction device 20 to maintain the experimental temperature within a predetermined experimental temperature range.
  • the state sensing device 50 is also used to collect experimental temperature data of the reaction device 20.
  • the main control system 60 is configured to: after receiving the experimental temperature data, parse the experimental temperature data to obtain a current experimental temperature, determine whether the current experimental temperature exceeds the predetermined experimental temperature range, and if so, generate a temperature reduction control instruction Or temperature rise control command.
  • the drive device 70 After receiving the temperature decrease control command or the temperature increase control command, the drive device 70 controls the temperature device to decrease or rise so that the current experimental temperature is maintained within a predetermined experimental temperature range.
  • the biochemical detection device for thioredoxin reductase (TR) detection provided in the seventh embodiment of the present invention further includes: a light source system 90 for providing predetermined experimental illumination to the cuvette in the reaction device 20 condition.
  • the invention can adopt the existing illumination adjustment technology, and details are not described herein again.
  • the biochemical detection device for thioredoxin reductase (TR) detection provided in the eighth embodiment of the present invention further includes: a power supply system 100, the main control system 60, the driving device 70, and the light source system 90 is connected to supply power to the main control system 60, the driving device 70, and the light source system 90.
  • the power system 100 includes a first power subsystem 101, a second power subsystem 102, a third power subsystem 103, and a fourth power subsystem 104.
  • the first power subsystem 101 is connected in series with the main control system 60 and the driving device 70 to supply power to the main control system 60 and the driving device 70; the second power subsystem 102 and the driving device 70 Connecting the power supply device 70 in series; the third power supply subsystem 103 is connected in series with the driving device 70 and the temperature control device 80 to supply power to the driving device 70 and the temperature control device 80; Subsystem 104, in series with the light source system 90, supplies power to the light source system 90.
  • the first power subsystem 101 is 5V DC
  • the second power subsystem 102 is 24V DC
  • the third source subsystem 33 and the fourth power subsystem 104 are 12V DC
  • the first power source The subsystem 101, the second power subsystem 102, the third power subsystem 103, and the fourth power subsystem 104 are connected in parallel.
  • the biochemical detection device for thioredoxin reductase (TR) detection provided in the ninth embodiment of the present invention further includes: a filter 4 electrically connected to the main control system 60 for a preset range of the input The internal alternating current is filtered, and the filtered alternating current is sent to the power supply system 100.
  • the alternating current in the preset range is 120V-250V.
  • Fig. 10 is a flow chart showing the operation method of the biochemical detecting device for detecting thioredoxin reductase (TR) according to the tenth embodiment of the present invention.
  • a method for operating a biochemical detection device for detecting thioredoxin reductase activity comprises:
  • Step S110 when the state sensing device detects that the target reagent and/or the target sample in the accommodating device is rotated to the target sampling hole and the reagent state information is the reagent addition completion state, generating a reagent collection instruction;
  • step S120 the driving device controls the sampling device to rotate to the target sampling hole of the loading device to collect the target reagent and/or the target sample based on the received reagent collection instruction.
  • Step S130 when the state sensing device detects that the target reaction cup in the reaction device rotates to the target liquid adding hole and the reagent state information of the sampling device is the reagent collecting completion state, generating a reagent adding instruction;
  • Step S140 the driving device controls the sampling device to rotate to the target liquid adding hole of the reaction device based on the received reagent adding instruction, and adds the target reagent and/or the target sample to the target reaction cup corresponding to the target liquid adding hole.
  • step S110 and step S120 are reagent addition processes.
  • step S130 and step S140 are reagent collection processes. Usually, the reagent is first collected and then the reagent is added, so step S120 is preceded by step S140.
  • the reagents are collected first, and the reagents are added after.
  • the signal that triggers the reagent acquisition is simply the signal that the target reagent and/or target sample is rotated to the target sampling well.
  • the signal that triggers the reagent collection is not only the signal that the target reagent and/or the target sample is rotated to the target sampling hole, but also the reagent addition completion state signal.
  • Fig. 11 is a flow chart showing the operation method of the biochemical detecting device for detecting thioredoxin reductase (TR) according to the eleventh embodiment of the present invention.
  • the method for operating the biochemical detection device for detecting thioredoxin reductase (TR) according to the eleventh embodiment of the present invention further includes:
  • Step S150 when the state sensing device detects that the reagent state information of the sampling device is that the reagent addition is completed, generating a stirring command;
  • step S160 the driving device controls the stirring device to rotate to the target liquid adding hole to stir the mixed liquid in the target reaction cup based on the received stirring command.
  • Figure 12 is a flow chart showing the operation method of the biochemical detection device for thioredoxin reductase (TR) detection according to the twelfth embodiment of the present invention.
  • the method for operating a biochemical detection device for detecting thioredoxin reductase (TR) according to the eleventh embodiment of the present invention further includes:
  • Step S170 when the state sensing device detects that the stirring state information of the stirring device is that the stirring is completed, generating a stirring reset command;
  • step S180 the driving device controls the stirring device to reset based on the received agitation reset command.
  • Figure 13 is a flow chart showing the operation method of the biochemical detection device for thioredoxin reductase (TR) detection according to the thirteenth embodiment of the present invention.
  • the method for operating a biochemical detection device for detecting thioredoxin reductase (TR) according to the thirteenth embodiment of the present invention further includes:
  • Step S210 when the state sensing device detects that the number of times the reaction device rotates by a predetermined angle is a predetermined number of times, generating a reagent rotation command
  • step S220 the driving device controls the accommodating device to rotate by a predetermined angle based on the received reagent rotation command, so that the next target cuvette is aligned with the target liquid adding hole.
  • the method further comprises:
  • the cleaning device controls the cleaning device to clean the target reaction cup corresponding to the target liquid filling hole based on the received cleaning instruction.
  • the number of cuvettes on the reaction disk is 81, which are divided into a control group and an experimental group.
  • the number of cuvettes in the experimental group was 40.
  • the 81 cuvettes are divided into 9 groups, each of which includes 9 cuvettes.
  • the accommodating disc has three rings to accommodate the fixing members, and the number of fixing members per lap is 40.
  • the first ring receiving fixing member is for holding the first reagent bottle
  • the second ring receiving fixing member is for holding the sample bottle
  • the third ring receiving fixing member is for holding the second reagent bottle.
  • the ultimate goal of the present invention is to add the first reagent, the sample, and the second reagent to all of the 81 cuvettes.
  • the first reagent, the second reagent and the sample involved in the examples of the present invention are all reagents related to the detection of thioredoxin reductase activity.
  • the sample is blood, body fluid or tissue homogenate.
  • the first reagent includes a working solution (a mixture solution of TrisHCL, morpholinylpropanesulfonic acid, disodium hydrogen phosphate citrate buffer system, and disodium hydrogen phosphate disodium hydrogen phosphate buffer solution) and a thioredoxin reductase inhibitor compound. .
  • the second reagent is a mixed reagent comprising 5,5-dithiobis(2-nitrobenzoic acid) or substituted 6,6'-dinitro-3,3'-dithiobenzoic acid and nicotinamide adenine A mixed solution of dinucleotide phosphoric acid.
  • the working fluid configuration process is: according to the ratio of 1:1:2:4, the reagent TrisHCL (0.025-0.125 mol / L, PH 5.5.8-7.2), morpholinyl C Sulfonic acid (0.25mol/L), disodium hydrogen phosphate citrate buffer system 0.2mol/L, and sodium hydrogen phosphate disodium phosphate potassium dihydrogen phosphate buffer solution 1-15mol/L; wherein the hydrogen phosphate disodium citrate buffer The pH of the system is 2.2-8.0; the pH of the potassium hydrogen phosphate disodium phosphate buffer solution is 4.9-8.2; then the TrisHCL (trishydroxymethylaminomethane hydrochloride), morpholinyl propanesulfonic acid The disodium hydrogen phosphate citrate buffer system and the potassium hydrogen phosphate disodium hydrogen phosphate buffer solution are uniformly mixed.
  • the working solution and the inhibitor are mixed in a ratio of 1:1 to 1:5 to form the inhibitor solution; the inhibitor solution is uniformly mixed; wherein the inhibitor is thioredoxin reductase inhibition Compound. It can be a chemical monomer selenium compound.
  • the mixed reagent generated by the mixing is highly accurate at the time of detection, wherein the A reagent is 5,5-dithiobis (2- Nitrobenzoic acid) or substituted 6,6'-dinitro-3,3'-dithiobenzoic acid; B reagent is nicotinamide adenine dinucleotide phosphate.
  • a reagent is 5,5-dithiobis (2- Nitrobenzoic acid) or substituted 6,6'-dinitro-3,3'-dithiobenzoic acid
  • B reagent is nicotinamide adenine dinucleotide phosphate.
  • the present invention provides three liquid sampling holes, and the first reagent corresponds to the first liquid sampling hole, and the sample corresponds to the first The second liquid collecting hole corresponds to the third liquid collecting hole.
  • the present application provides two liquid addition holes (the first liquid addition hole and the second liquid addition hole).
  • the first cycle group only the first reagent is added to the first reaction cup, and the first reagent may be added through the first liquid addition hole or the second liquid addition hole.
  • the second cycle group a sample is added to the first set of reaction cups, and a first reagent is added to the second set of reaction cups, and the first reagent and the sample can be added through the first liquid addition hole and the second liquid addition hole.
  • a second reagent is added to the first reaction cup, a sample is added to the second reaction cup, and a first reagent is added to the third reaction cup, and then, according to the liquid addition sequence of the third group, continue to The first set of reagents, the sample and the second reagent are added to the second set of cuvettes to the ninth set of reaction cups.
  • the driving device controls the rotation of the accommodating disk, and when the state detecting device 50 detects that the first reagent in the accommodating device rotates to the first liquid filling hole or the second liquid adding hole, and the reagent state information When the reagent is added to the completion state, a first reagent collection instruction is generated;
  • the driving device controls the sampling device to rotate to the first sampling hole of the holding device to collect the first reagent based on the received first reagent collecting instruction;
  • the driving device 70 controls the sampling device 30 to rotate to the first liquid filling hole or the second liquid adding hole of the reaction device 20 and add the first reagent to the first liquid filling hole or the first The second addition liquid hole corresponds to the first reaction cup.
  • the driving device controls the reaction device 20 to rotate, and when the state sensing device 50 detects that the first cuvette in the second group of reaction cups in the reaction device rotates to the first liquid addition hole
  • the reagent state information of the sampling device is the reagent collection completion state
  • a first reagent addition command is generated.
  • the driving device controls the sampling device to rotate to the first liquid adding hole of the reaction device based on the received reagent adding instruction, and adds the first reagent to the first reaction cup corresponding to the first liquid adding hole;
  • the driving device controls the sampling device to rotate to the first sampling hole of the receiving device to collect the first reagent based on the received first reagent collecting instruction;
  • the driving device 70 controls the sampling device 30 to rotate to the first liquid adding hole of the reaction device 20 and adds the first reagent to the first reaction cup corresponding to the first liquid filling hole.
  • the sampling device collects the sample according to the process of collecting the first reagent, adds the first reagent in the first cycle of the second cycle group, and adds the sample in the second cycle until the nine reaction cups in the second reaction cup are added.
  • the first reagent, and the nine cuvettes in the first set of cuvettes are added to the sample, and the second cycle group ends.
  • a first reagent is added to the third set of reaction cups; in the second cycle, a sample is added to the second set of reaction cups; and in the third cycle, a second reaction is added to the first set of reaction cups. Reagents until 9 cuvettes in the third set of cuvettes are added to the first reagent, 9 cuvettes in the second set of cuvettes are added to the sample reagent, and 9 cuvettes in the first set of cuvettes are added to the second reagent
  • the third cycle group ends.
  • the first reagent, the sample and the third reagent are added to the reaction cups of Groups 4-9 until the whole plate detection is completed or the detection is stopped.
  • reaction time is up to 22 cycles (22.5 s per cycle), preferably, For 20 cycles, for the set of cuvettes, if the set of reaction cups is rotated to the dosing position during the 22 cycles, the pipetting device is left in a waiting state.
  • the hydraulic device can be controlled by the client, and then the cleaning device can be controlled to remove the waste liquid in the reaction cup, and the reaction cup is cleaned. After the cleaning is completed, the reaction cup is The operation of adding the first reagent, the sample, and the second reagent can be continued.
  • the mixture comprising the first reagent, the sample and the second reagent in the first set of reaction cups begins to react, at this time, in the next cycle. If the first set of reaction cups of the reaction disk are rotated to the dosing position, the pipetting device is left in a waiting state.
  • biochemical detection device of the invention can not only detect biochemical reactions, but also detect non-biochemical reactions, it is only necessary to input biochemical reaction parameters or non-biochemical reaction parameters to be input on the user input interface of the client. Automatic adjustment according to the specific biochemical reaction or non-biochemical reaction detection process.
  • the present invention aims to provide a method for detecting thioredoxin reductase activity, which is capable of conforming to operation by setting a sample/reagent volume suitable for a synergistic detecting device for thioredoxin reductase activity described above.
  • the method for selecting a working fluid requires the method of the present invention to be used on a synergistic detecting device for thioredoxin reductase activity described above, including a method for loading a sample, a method for protecting from a light operation, and a method for operating a reagent mixture.
  • Such as the intelligent instruction method for the drive system is provided.
  • the intelligent instructions such as the number of cycles in each cycle group and the time of each cycle in the operation of the collaborative detection device, and the requirements for the operation process are all detection methods that are connected with the human peripheral blood TR function detection method.
  • the method of the present invention is a method for realizing the requirement of TR enzymatic detection function by using a synergistic detecting device for thioredoxin reductase activity described above, and is used for the synergistic detection of a thioredoxin reductase activity described above.
  • a method for detecting thioredoxin reductase activity of a device is a method for realizing the requirement of TR enzymatic detection function by using a synergistic detecting device for thioredoxin reductase activity described above, and is used for the synergistic detection of a thioredoxin reductase activity described above.
  • Another object of the present invention is also to protect a biochemical detection device for biochemical detection of thioredoxin reductase in human blood and an operation method thereof.
  • the biochemical detection device of the present invention realizes sulfur in human blood for the first time.
  • the fully automated detection of oxyredoxin reductase activity solves the problem of intelligent detection of thioredoxin reductase activity by manual in the prior art; on the other hand, the biochemical detection device of the present application can not only realize a single sample set TR The inspection process also enables multiple samples to be continuously completed in the TR detection process.
  • the completion of the overall detection process for a single sample can be achieved. Due to the continuous processing of the sample, the continuous loading and reaction time are strictly required. Since the sample disk rotates two squares, the reaction disk rotates one space, that is, two samples are taken from each sample tube, and two reaction cups are respectively added as an experimental group. And the detection of the data of the control group, so that the simultaneous detection of a single sample can be completed.
  • the hardware settings include the configuration of the reaction disk, the configuration of the sample disk, the linkage mode between the two disks, the sample loading and sampling device, etc., which can realize the completion of the multi-sample continuous detection process.
  • the improvement of the method includes the timing of the rotation time, the rotation interval, the rotation distance, the sampling sequence, the loading time and the like for the sample tray and the reaction tray. Since each reaction cup needs to add three times of solution, it can be reacted in each group. During the process of adding the solution to the cup, the addition of the next set of reaction cups is started at the same time, so that a plurality of samples can be continuously detected and can be cyclically detected, thereby shortening the detection time.
  • the present invention employs specific driving hardware, enabling software and hardware to work together, which can improve clinical TR detection efficiency and save costs.
  • the experimental results detected by the biochemical testing device of the present invention can meet the requirements of national testing standards.

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Abstract

一种硫氧还蛋白还原酶活性检测方法、检测设备及其操作方法，该检测方法包括配液，配置工作液、抑制剂溶液和混合试剂；加样本，将工作液加入对照组反应杯，将抑制剂溶液加入实验组反应杯，向对照组反应杯和实验组反应杯分别加入样本；温育，将对照组反应杯和实验组反应杯放置在预定温度的避光环境下孵育第一预定时间；测定，分别向对照组反应杯和实验组反应杯加入混合试剂，在预定波长时测定吸光度值第二预定时间。该检测方法和检测设备能够实现硫氧还蛋白还原酶活性的全自动化检测，快速高效，省时省力。

Description

一种硫氧还蛋白还原酶活性检测方法、检测设备及其操作方法 技术领域
本发明属于酶活性检测领域，具体涉及一种人血中硫氧还蛋白还原酶(TR)活性的生化检测方法、检测设备及其操作方法。
背景技术
硫氧还蛋白还原酶(TR)测定项目为国内外首创的临床肿瘤检测项目。该项目可以填补异常增生类疾病诊断缺乏临床检验方法的空白。
TR检测设备能够应用于体检人群的肿瘤早期筛查，住院人群的肿瘤疗效监测，复发预警及健康管理等，具有极大的市场需求和发展潜力。
现有技术《专利ZL201080049877.X：用于测定样品中硫氧还蛋白还原酶活性的方法和试剂盒及应用》是基于人工分步逐步操作实现，如：手工操作酶标仪、手工加样、手工操作避光、手工放置摇床实现摇匀，手工加样等；其用样本的各种条件也是针对手工操作过程和用量设计。由于人工手工操作包括了人工加样误差大；各种操作非连贯，容易产生中间误差；操作时间长不利于大规模临床检测应用等多种需要改善的问题。
现有技术中的TR检测试剂盒为初代研发产品，其检测步骤，动作，通量等方面都具有一定局限性，因此在检测速度和检测精度方面仍有改进和修正的空间。例如，只能通过手动检测人血中硫氧还蛋白还原酶(TR)，检测效率低；通过现有技术中的生化检测设备检测出的数据，不符合国家数据指标分布；等等。
因此，急需要提出一种全自动化的硫氧还蛋白还原酶(TR)检测的生化检测方法及检测设备，来提高检测效率且检测出的数据能够符合国家数据指标分布标准。
发明内容
本发明的目的是提供一种新的全自动化的硫氧还蛋白还原酶活性检测方法、检测设备及其操作方法，以节省检测时间和检测步骤。解决了现有技术中手工完成人血中硫氧还蛋白还原酶检测的问题，且能够提高临床中硫氧还蛋白还原酶检测的检测效率、节省成本。本发明首次实现了实现了“TR活性检测”在生化检测设备上的全自动化运行；通过本发明直接获得的数据符合国家检测标准要求，并能保证检测数据结果体现该检测技术的预警功能(符合国家数据指标分布)。
根据本发明的一个方面，提供了硫氧还蛋白还原酶活性检测方法，包括：配液，配置工作液、抑制剂溶液和混合试剂；加样本，将50uL-70uL的工作液加入对照组反应杯；将50uL-70uL抑制剂溶液加入实验组反应杯；向对照组反应杯和实验组反应杯分别加入10uL-30uL的样本；本加样量是根据自动化的协同检测设备最佳工作所需要体积量设计。样本温育，在避光环境下，将对照组反应杯和实验组反应杯放置在30℃-40℃温度下孵育第一预定时
间；测定，分别向对照组反应杯和实验组反应杯加入100uL-150uL的混合试剂；在预定波长下，测定第二预定时间段内的吸光度值。
进一步，配置工作液的步骤包括：按照1:1:2:4的比例取三羟甲基氨基甲烷盐酸盐、吗啉基丙磺酸、磷酸氢二钠柠檬酸缓冲体系和磷酸氢二钠磷酸二氢钾缓冲溶液；将自动化的协同检测设备三羟甲基氨基甲烷盐酸盐、吗啉 基丙磺酸、磷酸氢二钠柠檬酸缓冲体系和磷酸氢二钠磷酸二氢钾缓冲溶液混合均匀。
进一步，自动化的协同检测设备三羟甲基氨基甲烷盐酸盐的PH为5.5-7.2，浓度为0.025-0.125mol/L；自动化的协同检测设备吗啉基丙磺酸的浓度为0.25mol/L；自动化的协同检测设备磷酸氢二钠柠檬酸缓冲体系的PH为2.2-8.0，浓度为0.2mol/L；自动化的协同检测设备磷酸氢二钠磷酸二氢钾缓冲溶液的PH为4.9-8.2，浓度为1-15mol/L。
进一步，自动化的协同检测设备配置抑制剂溶液的步骤包括：以1:1-1:5的比例混合自动化的协同检测设备工作液和抑制剂，形成自动化的协同检测设备抑制剂溶液；将自动化的协同检测设备抑制剂溶液混合均匀，自动化的协同检测设备抑制剂为硫氧还蛋白还原酶抑制剂化合物。
进一步，自动化的协同检测设备配置混合试剂的步骤包括：以1:4-1:8的比例混合自动化的协同检测设备试剂A和自动化的协同检测设备试剂B，形成自动化的协同检测设备混合试剂；将自动化的协同检测设备混合试剂混合均匀；自动化的协同检测设备A试剂为5,5-二硫代双(2-硝基苯甲酸)或取代6,6'-二硝基-3,3'-二硫代苯甲酸；自动化的协同检测设备B试剂为烟酰胺腺嘌呤二核苷酸磷酸。
进一步，自动化的协同检测设备预定温度为30-40℃。
进一步，自动化的协同检测设备第一预定时间为8-20分钟。
进一步，自动化的协同检测设备第一预定时间为10分钟。
进一步，自动化的协同检测设备预定波长为405nm-450nm。
进一步，自动化的协同检测设备第二预定时间为20-30个周期。
本发明提供的一种人外周血硫氧还蛋白还原酶活性的检测方法，通过将试剂A和试剂B在自动化的协同检测设备上自动取样混合成混合试剂，经协同检测设备自动加入样本中进行混合、搅拌的操作，替代了分别添加试剂A和试剂B做手工重复混合、搅拌的工作，提高了工作效率。该检测方法通过对适用于上述介绍的一种硫氧还蛋白还原酶活性的协同检测设备的样本/试剂体积的设定，能够符合运行的工作液体选择的方法要求，本发明的方法在上述介绍的一种硫氧还蛋白还原酶活性的协同检测设备上使用时，包括加样操作方法、避光操作要求方法、试剂混合的操作要求方法等针对驱动系统的智能指令方法规定。其中的智能指令如协同检测设备运行中每一周期组中周期数以及每一周期时间的规定，针对运行过程要求等都是和实现人外周血TR功能检测方法衔接的检测方法学。本发明的方法是适用于上述介绍的一种硫氧还蛋白还原酶活性的协同检测设备实现TR酶学检测功能要求的方法，用于上述介绍的一种硫氧还蛋白还原酶活性的协同检测设备的硫氧还蛋白还原酶活性检测方法。
根据本发明的另一方面，提供了一种用于硫氧还蛋白还原酶检测的生化检测设备，包括：
容纳装置，用于盛放多个试剂和/或样本，并在驱动装置的驱动作用下，围绕轴心周期性转动使得目标试剂和/或目标样本转动至目标加液孔；
反应装置，用于盛放多个反应杯，并在驱动装置的驱动作用下，围绕轴心周期性转动使得目标反应杯转动至目标采样孔；
采样装置，用于在驱动装置的驱动作用下，围绕轴心周期性转动，将从目标采样孔中采集的所述目标试剂和/或目标样本加入目标加液孔对应的所 述目标反应杯中；
状态感应装置，用于检测所述反应装置的反应转动状态信息、容纳装置的容纳转动状态信息、采样装置的试剂状态信息；
主控系统，分别与所述状态感应装置、驱动装置连接，用于基于反应转动状态信息、容纳转动状态信息和试剂状态信息，生成并向驱动装置发送相应的控制指令；
驱动装置，分别与所述采样装置、所述容纳装置、所述反应装置连接，用于基于接收到的控制指令，控制所述采样装置、所述反应装置、所述容纳装置执行相应的操作。
根据本发明的另一个方面，提供了一种生化检测设备的操作方法，包括：
当状态感应装置检测到反应装置中目标反应杯转动至目标加液孔且采样装置的试剂状态信息为试剂采集完成状态时，生成试剂添加指令；
驱动装置基于接收到的试剂添加指令，控制采样装置转动至反应装置的目标加液孔并将目标试剂和/或目标样本添加到目标加液孔对应的目标反应杯中；
当状态感应装置检测到容纳装置中目标试剂和/或目标样本转动至目标采样孔时且试剂状态信息为试剂添加完成状态时，生成试剂采集指令；
驱动装置基于接收到的试剂采集指令，控制采样装置转动至盛放装置的目标采样孔处采集目标试剂和/或目标样本。
如上所述，本发明提出了一种全自动的用于人血中硫氧还蛋白还原酶(TR)检测的生化检测设备，实现了生化检测设备的全自动化，提高了检测效率、节省了成本。
附图说明
图1是本发明的硫氧还蛋白还原酶活性检测方法的流程示意图；
图2是本发明的硫氧还蛋白还原酶活性检测方法对应检测设备的结构示意图；
图3是本发明具体实施例的硫氧还蛋白还原酶活性检测方法的流程示意图；
图4是本发明具体实施例的硫氧还蛋白还原酶活性检测方法的又一流程示意图；
图5是图4所示流程图中步骤S1的流程图；
图6是图4所示流程图中步骤S2的流程图；
图7是图4所示流程图中步骤S3的流程图；
图8是本发明的用于硫氧还蛋白还原酶(TR)检测的生化检测设备的系统架构示意图；
图9是本发明的用于硫氧还蛋白还原酶(TR)检测的生化检测设备的机械结构示意图；
图10是本发明实施例十的硫氧还蛋白还原酶(TR)检测的生化检测设备的操作方法流程示意图；
图11是本发明实施例十一的硫氧还蛋白还原酶(TR)检测的生化检测设备的操作方法流程示意图；
图12是本发明实施例十二的用于硫氧还蛋白还原酶(TR)检测的生化检测设备的操作方法流程示意图；
图13是本发明实施例十三的用于硫氧还蛋白还原酶(TR)检测的生化检测设备的操作方法流程示意图；
图14是本发明一具体实例检测设备的检测原理示意图。
10.容纳装置，11.容纳盘，12.容纳固定件，20.反应装置，21.反应盘，22.反应杯固定件，30.采样装置，31.采样旋转件，32.采样固定件，33.采样针，40.搅拌装置，41.搅拌旋转件，42.搅拌固定件，43.搅拌针，50.状态感应装置，60.主控系统，70.驱动装置，80.温控装置，90.清洗装置，91.液压装置，92.清洗组件，100.电源系统100，101.第一电源子系统101，102.第二电源子系统，103.第三电源子系统，104.第四电源子系统，110.客户端。
具体实施方式
为使本发明的目的、技术方案和优点更加清楚明了，下面结合具体实施方式，对本发明进一步详细说明。应该理解，这些描述只是示例性的，而并非要限制本发明的范围。此外，在以下说明中，省略了对公知结构和技术的描述，以避免不必要地混淆本发明的概念。
现有技术中“TR活性检测试剂盒”所采用的检测方法(以下简称初代检测方法)有三个的技术局限，因而无法适用于自动化的协同检测设备：
(1)耗时较长，检测效率较低。初代的检测方法允许同时完成8-12个样本的检测，完成时间约为1.5-2个小时。而硫氧还蛋白酶活性的协同检测设备要求每40-50个样本的检测要在1.5个小时内完成，否则临床样本会因为在仪器中放置时间过长而失效；因此初代检测方法在面对大规模，高通量的TR临床自动化检测中无法适用；
(2)检测步骤繁多。初代检测方法包括7个步骤和动作，这对于检测人员的人工检测并不困难，但对于自动化的协同检测设备来说过于繁琐，因此会显著增加设备的动作时间，降低检测效率；
(3)初代检测方法在操作过程中需要两名检测人员相互配合操作，而自动化的协同检测设备的设计目的是希望单一检测人员即可完整操作仪器，因此两者的需求也并不相符。
在详述本发明实施方案前，需说明的是，本发明中的试剂A为用于检测硫氧还蛋白还原酶活性的试剂，试剂B为用于检测硫氧还蛋白还原酶活性的试剂，工作液通常为缓冲液，主要用于检测硫氧还蛋白还原酶活性的缓冲液。
本发明中的试剂A、试剂B均为已经通过了国家食品药品监督管理局的专家认证和审核获得了医疗器械注册证(证号为鄂食药监械(准)字2013第2401815号及国食药监械(准)字2014第3400264)的《硫氧还蛋白还原酶(TR)活性检测试剂盒》中的试剂A和试剂B，本发明中的工作液为上述《硫氧还蛋白还原酶(TR)活性检测试剂盒》中的试剂D，本发明中抑制剂为上述《硫氧还蛋白还原酶(TR)活性检测试剂盒》中的试剂C。
硫氧还蛋白还原酶(TR)是一种还原型辅酶II(NADPH)依赖的包含黄素腺嘌呤二核苷酸(FAD)结构域的二聚体硒酶，它和硫氧还蛋白、还原性辅酶II共同构成硫氧还蛋白系统。硫氧还蛋白还原酶在增生异常活跃的细胞中过量表达，该酶有启动细胞异常增生、启动凋亡抑制系统等与形成肿瘤密不可分的生理功能。TR活性与肿瘤的异常增生程度高度相关。因此，对硫氧还蛋白还原酶检测对肿瘤有重要的作用。
图1是本发明的硫氧还蛋白还原酶活性检测方法的流程示意图。
如图1所示，硫氧还蛋白还原酶活性检测方法包括：
步骤S1,配液，配置工作液、抑制剂溶液和混合试剂。
具体的，工作液通常为缓冲液，该工作液并不特别限定其浓度，优选的 配制成1X工作液浓度。
配制抑制剂溶液和混合试剂的步骤如下：
配置1.67mg/mL的试剂A；
配置10.29mg/mL的试剂B；
配置工作液；
其中，工作液配置过程为：按照1:1:2:4的比例取试剂三羟甲基氨基甲烷盐酸盐TrisHCL(0.025-0.125mol/L,PH 5.5-7.2)，吗啉基丙磺酸(0.25mol/L)，磷酸氢二钠柠檬酸缓冲体系0.2mol/L，和磷酸氢二钠磷酸二氢钾缓冲溶液1-15mol/L；其中，磷酸氢二钠柠檬酸缓冲体系的PH为2.2-8.0；磷酸氢二钠磷酸二氢钾缓冲溶液的PH为4.9-8.2；然后将TrisHCL(三羟甲基氨基甲烷盐酸盐)、吗啉基丙磺酸、磷酸氢二钠柠檬酸缓冲体系和磷酸氢二钠磷酸二氢钾缓冲溶液混合均匀。
配置抑制剂溶液；
具体的，以1:1-1:5的比例混合工作液和抑制剂，形成抑制剂溶液；将抑制剂溶液混合均匀；其中抑制剂为硫氧还蛋白还原酶抑制剂化合物。可以为化学单体硒啉类化合物。
其中，工作液和抑制剂混合的比例优选地为1：3的比例,在该比例下混合的工作液与抑制剂，是最经济的比例组合，即能对后续人外周血硫还蛋白还原酶活性检测更为准确，也是最为节约各试剂的组合。
以1:2-1:8的比例混合试剂A和试剂B，形成混合试剂；具体的，试剂A和试剂B的混合比例范围为1:2-1:8，优选的以1:4-5的比例来混合试剂A和试剂B，此时混合生成的混合试剂在检测时的准确度较高，其中，A试剂为 5,5-二硫代双(2-硝基苯甲酸)或取代6,6'-二硝基-3,3'-二硫代苯甲酸；B试剂为烟酰胺腺嘌呤二核苷酸磷酸。
将工作液混合均匀；将抑制剂溶液混合均匀；将混合试剂混合均匀。具体的，将混合均匀后的工作液、抑制剂溶液、混合试剂分别进行放置，通常放置在试剂槽中,以方便之后的检测。
步骤S2，加样本，将50uL-70uL的工作液加入对照组反应杯；将50uL-70uL抑制剂溶液加入实验组反应杯；向对照组反应杯和实验组反应杯分别加入10uL-30uL的样本；对照组反应杯和实验组反应杯间隔设置，例如：奇数编号的反应杯为对照组反应杯，偶数编号的反应杯为实验组反应杯。
具体的，向对照组反应杯和实验组反应杯加入的工作液的量是等同的，一般情况下测定一个样本时，在对照组反应杯和实验组反应杯一般加入50uL-70uL的工作液，优选的，向对照组反应杯和实验组反应杯加入50-60uL的工作液。优选的，向对照组反应杯和实验组反应杯加入等量的工作液，以使对照组和实验组的检测数据有可比较性，能对检测后的数据进行计算。
在使用本发明对硫氧还蛋白还原酶活性进行检测时，一般是1-16个样本为一组进行测定，每组样本的加样时间为约10分钟，即27个周期，每周期22.5秒，这组样本加样完后才能之后，就可以对本组样本进行温育，然后另一组样本再继续进行加样。
步骤S3，温育，在避光环境下，将对照组反应杯和实验组反应杯放置在30℃-40℃温度下孵育第一预定时间；其中，预定温度为30℃-40℃；第一预定时间为8-20分钟，优选的为10分钟。
具体的，加入样本之后，将对照组反应杯和实验组反应杯放置在 30℃-40℃的温度下进行避光孵育为8-20分钟，优选的为10分钟，由于该检测方法可配合检测设备使用，在预定时间为10分钟时，可以实现在硫氧还蛋白还原酶活性的协同检测设备上的自动化检测。一组样本孵育时间到后，即可对本组样本进行测定。
步骤S4，测定，分别向对照组反应杯和实验组反应杯加入100uL-150uL的混合试剂；在预定波长下，测定第二预定时间段内的吸光度值。具体的，在对样本进行测定时，首先用加样针向对照组反应杯和实验组反应杯加入110uL-130uL的混合试剂，最优的，可在对照组反应杯和实验组反应杯加入120uL的混合试剂，在405nm-450nm的波长下连续测定样本的吸光度值7.5-11.25分钟，即20-30个周期。
本发明中将试剂A和试剂B进行混合后，进行一块搅拌，使其均匀混合后，在测定时，直接将混合试剂一块加入待测定的试剂中，实现了混合加样，比现有技术中单独对试剂A和试剂B分别搅拌，然后单独加样，省略了一部分的步骤，节省了检测的时间，且实现了在硫氧还蛋白还原酶活性的协同检测设备上自动化检测，提高了检测的效率。，其中，本发明中的样本是指来自生物的任何组织或分离自其中的部分，样本优选血液、体液、组织匀浆液，并最优选血液，其中血液可以是血清、血浆等组分。
本发明的检测方法可适用于“一种硫氧还蛋白酶活性检测设备”，如图2所示，该设备的结构具体为：
检测设备包括：
壳体1，
容纳装置2，用于容纳样本和试剂(包括工作液/抑制剂溶液和混合试 剂)，并在驱动系统(图中未示出)的驱动作用下，围绕轴心做周期性转动；其中，容纳装置2包括：样本试剂盘2-1和分别沿样本试剂盘2-1圆周方向均匀分布的样本管固定部件2-2、工作液/抑制剂溶液固定部件2-3和混合试剂固定部件2-4；样本管固定部件2-2可以是围绕样本试剂盘圆周方向设置的一圈具有若干个孔的环状结构部件，放置于样本试剂盘中能够置放样本管，例如：试管架，样本管固定部件上的孔优选为40个；工作液/抑制剂溶液固定部件2-3和混合试剂固定部件2-4的结构与样本管固定结构类似，分别用于置放工作液/抑制剂溶液瓶或混合试剂瓶，数量均优选为30或40个，每一个样本管固定部件2-2、每一个工作液/抑制剂溶液固定部件2-3和每一个混合试剂固定部件2-4均是以样本试剂盘的中心为圆心，且沿着样本试剂盘的半径由内至外依次分布。
反应装置3，在驱动系统的驱动作用下，围绕轴心做周期性转动；反应装置3包括：反应盘3-1和沿反应盘圆周方向均匀分布的多个反应杯固定部件3-2，反应杯固定部件3-2可以是围绕反应盘圆周方向设置的一圈具有若干个孔的环状结构部件，放置于反应盘中能够容纳或置放反应杯，例如：试管架，反应杯的数量为81个，可分为9组。
采样装置4，用于基于容纳装置2和反应装置3的周期性转动，从容纳装置2中采集试剂和/或样本放入反应装置3；采样装置4可选的为采样针；
驱动系统，与容纳装置2、反应装置3和采样装置4连接，用于控制容纳装置2、反应装置3和采样装置4执行相应的操作。
液压装置5，包括真空泵5-1、真空罐5-2和液路机构5-3，液路机构包括管道和设置于管道上的阀门，液路机构还与采样装置和自动清洗装置连接，
真空泵5-1，用于调节真空罐5-2内的气压，以使真空罐5-2内气压达到预设气压；
真空罐5-2，用于在预设气压下，控制液路机构动作，以对采样装置进行上升、下降和旋转的操作，和使液体进入或排出自动清洗装置。
自动清洗装置6，与液压装置5连接，用于基于液压装置5的控制，对反应装置3中的反应杯进行清洗；自动清洗装置6可以是清洗针，能够吸入液体至清洗针内部的容纳空间，进而释放至反应杯中对反应杯进行清洗，再将清洗后的废液从反应杯中吸走排出至废水池(图中未示出)，废水池可设置于检测设备外部。
搅拌装置7，与驱动系统连接，用于对试剂和样本形成的混合液进行搅拌，在各加样步骤完成后进行均匀搅拌。优选地，搅拌装置可以是搅拌针。
温控装置8，位于多个反应杯固定部件下方，用于控制反应杯固定部件内的反应杯的温度保持在设定的实验温度。具体地，温控装置8是温控槽，为槽状结构，使得81个反应杯能够均位于槽状结构内部，以保持反应过程中反应装置内部的环境温度保持在反应温度和孵育温度。
光电装置9，设置于壳体1上表面，用于控制光路和波长，为样本与试剂的反应过程提供光照，连续测定样本的吸光度值。光电装置为光电盒。
注射器10，设置于壳体1上表面。
其中，样本试剂盘上设置有样本试剂盘盖，以对样本圆盘进行封闭为TR活性检测提供实验环境；样本试剂盘盖上设置有样本采样孔、工作液/抑制剂溶液采样孔和混合试剂采样孔；反应盘上设置有第一加液孔和第二加液孔；样本采样孔、工作液/抑制剂溶液采样孔、混合试剂采样孔、第一加液孔和第 二加液孔均位于以采样装置为圆心的同一个圆上，采样装置沿着圆的圆周在样本采样孔、工作液/抑制剂溶液采样孔、混合试剂采样孔、第一加液孔和第二加液孔之间做周期性运动。
具体地，第一加液孔和第二加液孔之间的间隔可以设置为少于每组反应杯数量的数量个反应杯的距离，优选地，第一加液孔和第二加液孔之间的间隔可以设置为7-11个反应杯的距离，更为优选地，是设置为8个反应杯的距离。
本发明实施例还包括上位机，上位机与液压装置和驱动系统连接，用于向液压装置和驱动系统发送操作指令，以使得驱动系统控制容纳装置2和反应装置3做周期性转动，当需要采样时，驱动系统接收上位机的指令，控制容纳装置转动，容纳装置每转动一格，都会有一个样本管固定部件、工作液/抑制剂溶液瓶固定部件和混合试剂瓶固定部件转动至分别对应样本试剂盘盖上设置的样本采样孔、工作液/抑制剂溶液采样孔和混合试剂采样孔的位置处，采样装置会在液压装置的控制下下降至样本管固定部件、工作液/抑制剂溶液瓶固定部件和混合试剂瓶固定部件中，进而从样本管、工作液/抑制剂溶液瓶或混合试剂瓶中采集样本、工作液/抑制剂溶液或混合试剂然后在液压装置的控制下上升，再移动至第一加液孔或第二加液孔的位置处，在液压装置的作用下下降，将采集的样本、工作液/抑制剂溶液或混合试剂放入第一加液孔或第二加液孔位置处的反应杯中，与此同时，反应装置转动一格，使得第一加液孔和第二加液孔分别对应下一个需要加液的反应杯，进而继续完成下一个周期的采集以及加液。
容纳装置，用于容纳样本和试剂，并在驱动系统的驱动作用下，围绕轴 心做周期性转动；反应装置，用于容纳反应杯和实验杯，反应杯和实验杯在驱动系统的驱动作用下，围绕轴心做周期性转动；采样装置，用于基于容纳装置和反应装置的周期性转动，从容纳装置中采集试剂和/或样本放入反应装置；驱动系统，分别与容纳装置、反应装置和采样装置连接，用于控制容纳装置、反应装置和采样装置动作。
下面结合该检测设备对本发明的检测方法进行详细说明：
需说明的是，在使用本发明的方法对人外周血硫氧还蛋白还原酶的活性进行检测时，有时需要对照组和实验组两组进行检测，以便进行对照，检测本发明的准确性和快速性。
下面的方法即可适用于实验组也适用于对照组，其中检测方法包括：
图3和图4显示了本发明实施例的硫氧还蛋白还原酶活检测方法的流程示意图。
如图3和图4所示，一种硫氧还蛋白还原酶活检测方法，包括：
S1，从第一周期开始，驱动系统控制采样装置从样本试剂盘中依次采集工作液/抑制剂溶液，并控制采样装置将采集的工作液/抑制剂溶液依次加入第一组反应杯，直至第一周期组结束；
具体地，如图5所示，步骤S1包括：
S11，在第一周期组的第一个周期中，驱动系统控制反应盘转动，使得第一组反应杯中的第一个反应杯位于第一加液孔位置处；
S12，驱动系统控制采样装置转动至工作液/抑制剂溶液采样孔位置处，从样本试剂盘的某一个工作液/抑制剂溶液瓶中采集工作液/抑制剂溶液；
S13，驱动系统控制采样装置转动至第一加液孔位置处，将采集的工作液 /抑制剂溶液加入第一加液孔位置处的反应杯内；
S14，驱动系统控制采样装置和反应盘重复上述工作液/抑制剂溶液的加液步骤，直至第一组反应杯中的每一个反应杯均加入工作液/抑制剂溶液，第一周期组结束。
具体地，在第一周期组中，当反应盘转动两个预设角度时，样本试剂盘转动一个预设角度，直至第一周期组结束。优选地，每一个预设角度可以设置为两个反应杯固定部件的间隔，这样设置能够使得每转动一个预设角度就可以使得第一组反应杯中的每一个反应杯依序位于第一加液孔位置处；
S2，从第二周期组的第一个周期开始，每间隔两个周期，驱动系统控制采样装置从样本试剂盘中依次采集工作液/抑制剂溶液，并控制采样装置将采集的工作液/抑制剂溶液依次加入第二组反应杯；
从第二周期组的第二个周期开始，每间隔两个周期，驱动系统控制采样装置从样本试剂盘中依次采集样本，并依次控制采样装置将采集的样本加入第一组反应杯，直至第二周期组结束；
具体地，如图6所示，步骤S2包括：
S21，在第二周期组的第一个周期中，驱动系统控制反应盘转动，使第二组反应杯中的第一个反应杯位于第一加液孔位置处；
S22，驱动系统控制采样装置转动至工作液/抑制剂溶液采样孔位置处，从样本试剂盘的某一个工作液/抑制剂溶液瓶中采集工作液/抑制剂溶液，并控制采样装置转动至第一加液孔位置处，将采集的工作液/抑制剂溶液加入第一加液孔位置处的反应杯内；
S23，在第二周期组的第二个周期中，驱动系统控制反应盘转动，使第一 组反应杯中的第一个反应杯位于第二加液孔位置处；
S24，驱动系统控制采样装置转动至样本采样孔位置处，从样本试剂盘的某一个样本管中采集样本，并控制采样装置转动至第二加液孔位置处，将采集的样本加入第二加液孔位置处的反应杯内；
S25，在第二周期组的第三个周期中，采样装置不动作；
具体地，在第二周期组的第三个周期中，反应盘可以转动，也可以不转动，若第二周期组的第三个周期选择不转动，则可以选择在下一周期中转动，但为了考虑程序控制的准确度以及方便性，反应盘优选地为不转动；以及若此周期中，样本试剂盘需要转动，则也可以选择在本周期转动或者在下一周期转动。
S26，按照上述第二周期组的三个周期的加液步骤循环地加液，直至第一组反应杯中的每个反应杯均加入样本，且第二组反应杯中的每个反应杯均加入工作液/抑制剂溶液，第二周期组结束。
S3，从第三周期组的第一个周期开始，驱动系统每间隔两个周期控制采样装置从样本试剂盘中依次采集工作液/抑制剂溶液，并控制采样装置将采集的工作液/抑制剂溶液依次加入第三组反应杯；
从第三周期组的第二个周期开始，驱动系统每间隔两个周期控制采样装置从样本试剂盘中依次采集样本，并控制采样装置将采集的样本依次加入第二组反应杯；
从第三周期组的第三个周期开始，驱动系统每间隔两个周期控制采样装置从样本试剂盘中依次采集混合试剂，并控制采样装置将采集的混合试剂依次加入第一组反应杯，直至第三周期组结束。
第三周期组结束时，第一组反应杯即加液结束，开始进入孵育时间。
具体地，如图7所示，步骤S3包括：
S31，在第三周期组的第一个周期中，驱动系统控制反应盘转动，使得第三组反应杯中的第一个反应杯位于第一加液孔或第二加液孔位置处；
S32，驱动系统控制采样装置转动至工作液/抑制剂溶液采样孔位置处，从样本试剂盘的某一个工作液/抑制剂溶液瓶中采集工作液/抑制剂溶液，并控制采样装置转动至第一加液孔或第二加液孔位置处，将采集的工作液/抑制剂溶液加入第一加液孔或第二加液孔位置处的反应杯内；
S33，在第三周期组的第二个周期中，驱动系统控制反应盘转动，使得第二组反应杯中第第一个反应杯位于第一加液孔或第二加液孔位置处；
S34，驱动系统控制采样装置转动至样本采样孔位置处，从样本试剂盘的某一个样本管中采集样本，并控制采样装置转动至第一加液孔或第二加液孔位置处，将采集的样本加入位于第一加液孔或第二加液孔位置处的反应杯内；
S35，在第三周期组的第三个周期中，驱动系统控制反应盘转动，使得第一组反应杯中的第一个反应杯位于第一加液孔或第二加液孔位置处；
S36，驱动系统控制采样装置转动至混合试剂采样孔位置处，从样本试剂盘的某一个混合试剂瓶中采集混合试剂，并控制采样装置转动至第二加液孔位置处，将采集的混合试剂加入第一加液孔或第二加液孔位置处的反应杯内；
S37，按照上述第三周期组的三个周期的加液步骤循环在第一组反应杯、第二组反应杯和第三组反应杯中的每个反应杯之间进行加液，直至第一组反应杯均加入混合试剂，第二组反应杯中均加入样本，第三组反应杯均加入工作液/抑制剂溶液，第三周期组结束。
S4，按照第三周期组的加液步骤进行循环，将工作液/抑制剂溶液、样本和混合试剂分别并依次加入每组反应杯，直至整盘检测完成或停止检测；
具体地，按照第三周期组的三个周期的加液步骤循环加液，直至所有反应杯中均加入工作液/抑制剂溶液、样本和混合试剂或停止检测。
步骤S4在实施时，具体是从第四周期组的第一个周期开始，每间隔两个周期，依次采集工作液/抑制剂溶液，并依次加入第四组反应杯；从第四周期组的第二个周期开始，每间隔两个周期，依次采集样本，并依次加入第三组反应杯；从第四周期组的第三个周期开始，每间隔两个周期，依次采集混合试剂，并依次加入第二组反应杯；
对于反应装置来说，只要反应装置在工作，即可重复上述步骤S3，需要说明的是，待每一组反应杯都加入了工作液/抑制剂溶液、样本和混合试剂之后，即开始发生反应，反应时间最长为22个周期(每个周期22.5s)，优选地，为20个周期，对该组反应杯来说，在这22个周期内，若该组反应杯转动至加液位置，则采样装置静置处于等待状态。待该组反应杯反应结束后，可通过上位机发送指令控制液压装置，进而控制自动清洗装置将反应杯中反应后的废液抽走，并对反应杯进行清洗，清洗完成后该组反应杯即可继续进行加入工作液/抑制剂溶液、样本和混合试剂的操作。
例如：对于第一组反应杯来说，第三周期组结束后，第一组反应杯内的包括工作液/抑制剂溶液、样本和混合试剂的混合液即开始发生反应，此时，在下一个周期内，若反应盘的第一组反应杯转动至加液位置，则采样装置静置处于等待状态。
本发明实施例中协同检测设备执行相应的测试流程，81个反应杯分为9 组，每个反应共有74个测试周期，包括加入第一试剂至加入样本的27个周期，加入样本至加入第二试剂的27个周期，以及反应的20个周期，每个周期用时22.5秒，在本发明实施例中，
1)1-9周期(每个周期22.5s)：
在1-9周期的每个周期内，依次采集工作液/抑制剂溶液并依次加入1-9号反应杯中；
具体为：采样装置自工作液/抑制剂溶液采样孔位置处取工作液/抑制剂溶液(工作液或抑制剂溶液)，加入至反应装置的1号反应杯中，随即反应装置转动1格，花费1个周期(22.5s)；
连续重复上述动作共9次，将工作液/抑制剂溶液加入1-9号反应杯(此为第一组)中，共花费9个周期，反应杯转动9格；其中，反应装置每转动两次，容纳装置转动一次。即采样装置自样本试剂盘的每个工作液/抑制剂溶液瓶采集2次样本，然后容纳装置转动一次。
2)10-36周期(每个周期22.5s)：
在第10、13、16、19、22、25、28、31和34周期中，依次采集样本，并依次将样本加入1-9号反应杯；
在第11、14、17、20、23、26、29、32和35周期内，依次采集工作液/抑制剂溶液，并依次将工作液/抑制剂溶液加入10-18号反应杯；
在12、15、18、21、24、27、30、33和36周期内，采样装置不动作。
3)37周期开始：
从第37周期开始每间隔3个周期依次采集工作液/抑制剂溶液，并依次加入19-27号反应杯；从第38周期开始每间隔3个周期依次采集样本，并依次加入10-18号反应杯；从第39周期开始每间隔3个周期依次采集混合试剂， 并依次加入1-9号反应杯。
具体地，是在第37周期，加入工作液/抑制剂溶液至19号反应杯；第38周期，加入样本至10号反应杯；第39周期，加入混合试剂至1号反应杯；第40周期，加入工作液/抑制剂溶液至20号反应杯；第41周期，加入样本至11号反应杯；第42周期，加入混合试剂至2号反应杯，依此循环，至整盘检测完成或停止检测；
可选的，在整盘检测完成后，前面已经反应结束的反应杯组可在清洗后进入下一轮检测。
在整个检测过程中，反应盘可以设置为是顺时针转动；
当反应盘为顺时针转动时，第一周期组的每个周期中反应盘为顺时针转动，对于第一组反应杯中的第一个反应杯来说，可以通过顺时针或逆时针转动将其转至位于第一加液孔或第二加液孔位置处进行加液，之后的检测过程中，如涉及到待加液的反应杯无法按照每一周期转动一个预设角度转动至第一或第二加液孔位置处时，都可以通过顺时针转动转至两个加液孔位置处，可选的，还可以是通过逆时针转动。
通过上述加液步骤以及周期时间的设定，还有两盘之间转动的配置，使得两个盘协同工作，对于1号反应杯来说，加入样本至加入混合试剂等待27个周期(约为10min)，符合TR活性检测的医学要求；根据上述方法实现对人血样本检测，实现了通过TR特异性抑制剂对人血样本背景减扣，从而保证检测方法符合TR酶活性检测结果与国家相关标准的一致性(正常人群TR检测数据小于4单位，肿瘤高相关人群TR检测数据大于12单位)。
本发明硫氧还蛋白还原酶活性检测方法通过改进初代检测方法，可以满 足自动化检测的要求，进一步具有下述优点：
1)与已公开的TR活性检测技术方法(PCT/CN2010/078369)相比，显著减少孵育时间，单一样本的孵育时间从30min降低至10min，能够有效减少检测时间，使每40-50个样本的检测可以在1.5个小时内完成，实现协同检测设备的连续检测，满足硫氧还蛋白还原酶活性的协同检测设备的检测通量和速度要求。
2)与已公开的TR活性检测技术方法(PCT/CN2010/078369)相比，显著减少了检测的动作步骤，从7个动作步骤缩减至3-4个动作步骤，优化检测步骤，缩减了仪器的动作时间并方便仪器程序操作，使单个检测人员可以独立操控仪器，完成整个检测流程。
3)本发明的检测方法能够适应配合硫氧还蛋白还原酶活性的协同检测设备在TR临床自动化检测方面的动作要求，配置要求和指令要求；
4)经由本发明方法，在硫氧还蛋白还原酶活性的协同检测设备上完成临床TR活性检测，并经过特定软件(具体请见另一项申请专利：一种硫氧还蛋白还原酶活性的分析方法及系统)处理，能够成为临床医学认定的TR活性结果，并满足相应上市产品“TR活性检测试剂盒”及国家医疗器械注册产品标准的相关要求：YZB/国(Q/CVH 001-2011)；
5)本发明的检测设备和方法通过使用现有的“硫氧还蛋白还原酶(TR)活性检测试剂盒”(证号为鄂食药监械(准)字2013第2401815号及国食药监械(准)字2014第3400264)，所产生的TR检测结果满足相应上市产品“TR活性检测试剂盒”及国家医疗器械注册产品标准的相关要求：YZB/国(Q/CVH 001-2011)。
如上所述，本发明提供的硫氧还蛋白还原酶活性检测方法中，通过将试剂A和试剂B在上述介绍的一种硫氧还蛋白还原酶活性的协同检测设备上自动取样混合成混合试剂，经硫氧还蛋白还原酶活性的协同检测设备自动加入样本中进行混合、搅拌的操作，替代了分别添加试剂A和试剂B做手工重复混合、搅拌的工作，提高了工作效率。
本发明的另一目的是提供一种用于人血中硫氧还蛋白还原酶(TR)活性检测的生化检测设备，用于实现全自动的生化检测，提高检测效率、节省成本。
图8是本发明的用于硫氧还蛋白还原酶(TR)检测的生化检测设备的系统架构示意图。
如图8所示，本发明实施例一提供了一种生化检测设备，包括：容纳装置10，反应装置20，采样装置30，状态感应装置50，主控系统60，驱动装置70。
容纳装置10，用于盛放多个试剂和/或样本，并在驱动装置70的驱动作用下，围绕轴心周期性转动使得目标试剂和/或目标样本转动至目标加液孔。
反应装置20，用于盛放多个反应杯，并在驱动装置70的驱动作用下，围绕轴心周期性转动使得目标反应杯转动至目标采样孔。
采样装置30，用于在驱动装置70的驱动作用下，围绕轴心周期性转动，将从目标采样孔中采集的所述目标试剂和/或目标样本加入目标加液孔对应的所述目标反应杯中。
状态感应装置50，用于检测所述反应装置20的反应转动状态信息、容纳装置10的容纳转动状态信息、采样装置30的试剂状态信息。
主控系统60，分别与所述状态感应装置50、驱动装置70连接，用于基于反应转动状态信息、容纳转动状态信息和试剂状态信息，生成并向驱动装置70发送相应的控制指令；
驱动装置70，分别与所述采样装置30、所述容纳装置10、所述反应装置20连接，用于基于接收到的控制指令，控制所述采样装置30、所述反应装置20、所述容纳装置10执行相应的操作。
图9是本发明的用于硫氧还蛋白还原酶(TR)检测的生化检测设备的机械结构示意图。
容纳装置10包括：容纳盘11，活动设置，围绕轴心做周期性转动。容纳盘11上具有多个容纳固定件12，用于盛放试剂瓶和/或样本瓶。
可选的，在容纳盘11上设置至少一圈容纳固定件12，每圈具有多个容纳固定件12。容纳固定件12的多少可以根据用户的具体需要进行选择和设置。
优选的，可以设置多个容纳固定件12沿容纳盘11的边缘均匀设置，即每两个容纳固定件12之间均间隔预定距离，形成均匀设置的至少一圈容纳固定件12。
对于每个容纳固定件12，其既可以容纳试剂瓶座也可以容纳样本瓶座，当需要增加样本位减少试剂位时，可以将试剂瓶座取下，换上样本瓶座；当需要增加试剂位减少样本位时，则可以将样本瓶座取下而换上试剂瓶座，实现了试剂位和样本位的灵活互换，能够满足不同客户对样本位和试剂位的不同需求。
如图9所示，所述反应装置20包括：反应盘21，活动设置，围绕轴心 做周期性转动。多个反应杯固定件22，用于盛放反应杯。
可选的，在反应盘21上设置至少一圈反应杯固定件22，每圈具有多个反应杯固定件22。反应杯固定22的多少可以根据用户的具体需要进行选择和设置。
优选的，可以设置多个反应杯固定件22沿反应盘21的边缘均匀设置，即每两个反应杯固定件22之间均间隔预定距离，形成均匀设置的至少一圈反应杯固定件22。
如图9所示，采样装置30包括：采样旋转件31，活动设置，用于在驱动装置70的驱动作用下，围绕轴心周期性转动；采样固定件32，固定设置在采样转动件31上，在采样旋转件32的带动作用下转动；采样针33，一端为固定端，另一端为自由端。
结合图8-9，说明试剂添加过程：
主控系统60在接收到目标反应杯转动至目标加液孔时的反应转动状态信息且试剂状态信息为试剂采集完成状态时，生成试剂添加指令；所述驱动装置70在接收到试剂添加指令后，控制采样装置30转动至反应装置20的目标加液孔并将目标试剂和/或目标样本添加到目标加液孔对应的目标反应杯中。
结合图8-9，说明试剂采集过程：
主控系统60在接收到目标试剂和/或目标样本是否转动至目标采样孔时的容纳转动状态信息时和试剂状态信息为试剂添加完成状态时，生成试剂采集指令；所述驱动装置70在接收到试剂采集指令后，控制采样装置30转动至盛放装置10的目标采样孔处采集目标试剂和/或目标样本。
本发明实施例三提供的用于硫氧还蛋白还原酶(TR)检测的生化检测设备还包括：搅拌装置40，用于在驱动装置70的驱动作用下，围绕轴心做周期性转动以对目标反应杯中形成的混合液进行搅拌。
如图9所示，所述搅拌装置40包括：搅拌旋转件41，固定设置，用于在驱动装置70的驱动作用下，围绕轴心周期性转动；搅拌固定件42，固定设置在搅拌转动件31上，在搅拌旋转件42的带动作用下转动；搅拌针43，一端为固定端，另一端为自由端。
结合图8-9，说明搅拌过程：
主控系统60在接收到试剂状态信息为试剂添加完成时，生成搅拌控制指令；
驱动装置70在接收到搅拌控制指令后，控制搅拌装置40转动到目标加液孔以对目标反应杯中的混合液进行搅拌。
在本发明实施例四提供的用于硫氧还蛋白还原酶(TR)检测的生化检测设备中，所述状态感应装置50还用于检测搅拌装置40的搅拌状态信息；所述主控系统60，用于在接收到搅拌状态信息为搅拌完成后，生成搅拌复位指令；驱动装置，与搅拌装置40连接，控制搅拌装置40复位。
结合图8-9，说明容纳装置10的转动过程：
主控系统60在接收到转动预定角度的次数为预定次数时的反应转动状态信息时，生成试剂转动指令；容纳装置10在接收到试剂转动指令后，控制容纳装置10转动预定角度，使得下一个目标反应杯对准目标加液孔。
在本发明实施例五提供的用于硫氧还蛋白还原酶(TR)检测的生化检测设备中，还包括：清洗装置90，用于将清洗液中的液体输入所述目标加液孔 对应的目标反应杯以对所述目标反应杯进行清洗。
如图9所示，清洗装置90包括：液压装置91，用于控制清洗液中液体进入目标反应杯或将目标反应杯中的废液排出目标反应杯；清洗组件92，与所述液压装置91连接，在所述液压装置91的作用下，对目标反应杯进行清洗。
在本发明实施例六提供的用于硫氧还蛋白还原酶(TR)检测的生化检测设备中，还包括：
客户端110，与所述主控系统60连接，用于提供用户操作指令输入界面，并用于采集用户在所述用户操作指令输入界面输入的清洗指令，将所述清洗指令发送给所述主控系统60；所述驱动装置70在接收到所述清洗指令后，控制所述清洗装置90对目标加液孔对应的目标反应杯进行清洗。所述客户端110包括但不限于上位机。
在本发明实施例六提供的用于硫氧还蛋白还原酶(TR)检测的生化检测设备中，还包括：
温控装置80，在驱动装置70的驱动作用下，调节反应装置20的实验温度，使实验温度保持在预定实验温度范围。
结合图8-9，说明温度调节过程：
所述状态感应装置50还用于采集反应装置20的实验温度数据。
所述主控系统60，用于在接收到实验温度数据后，解析所述实验温度数据得到当前实验温度，判断所述当前实验温度是否超过所述预定实验温度范围，若是，生成温度降低控制指令或温度升高控制指令。
驱动装置70在接收到温度降低控制指令或温度升高控制指令后，控制温 度装置降低或升高，使得当前实验温度保持在预定实验温度范围。
本发明实施例七提供的用于硫氧还蛋白还原酶(TR)检测的生化检测设备中，还包括：光源系统90,用于为所述反应装置20中的所述反应杯提供预定实验光照条件。本发明可以采用现有的光照调节技术，此处不再赘述。
本发明实施例八提供的用于硫氧还蛋白还原酶(TR)检测的生化检测设备中，还包括：电源系统100，与所述主控系统60、所述驱动装置70、所述光源系统90连接，为所述主控系统60、所述驱动装置70、所述光源系统90供电。
所述电源系统100包括：第一电源子系统101、第二电源子系统102、第三电源子系统103、第四电源子系统104。
第一电源子系统101，与所述主控系统60、所述驱动装置70串联，为所述主控系统60、所述驱动装置70供电；第二电源子系统102，与所述驱动装置70串联，为所述驱动装置70供电；第三电源子系统103，与所述驱动装置70和所述温控装置80串联，为所述驱动装置70和所述温控装置80供电；第四电源子系统104，与所述光源系统90串联，为所述光源系统90供电。其中，所述第一电源子系统101为5V直流电，所述第二电源子系统102为24V直流电，所述第三源子系统33、第四电源子系统104为12V直流电；所述第一电源子系统101、第二电源子系统102、第三电源子系统103和第四电源子系统104之间为并联。
本发明实施例九提供的用于硫氧还蛋白还原酶(TR)检测的生化检测设备中，还包括：滤波器4，与所述主控系统60电连接，用于对输入的预设范围内的交流电进行滤波，将滤波后的交流电发送给所述电源系统100。其中， 所述预设范围内的交流电为120V-250V。
图10是本发明实施例十的硫氧还蛋白还原酶(TR)检测的生化检测设备的操作方法流程示意图。
如图10所示，一种用于硫氧还蛋白还原酶活性检测的生化检测设备的操作方法，包括：
步骤S110，当状态感应装置检测到容纳装置中目标试剂和/或目标样本转动至目标采样孔时且试剂状态信息为试剂添加完成状态时，生成试剂采集指令；
步骤S120，驱动装置基于接收到的试剂采集指令，控制采样装置转动至盛放装置的目标采样孔处采集目标试剂和/或目标样本。
步骤S130，当状态感应装置检测到反应装置中目标反应杯转动至目标加液孔且采样装置的试剂状态信息为试剂采集完成状态时，生成试剂添加指令；
步骤S140，驱动装置基于接收到的试剂添加指令，控制采样装置转动至反应装置的目标加液孔并将目标试剂和/或目标样本添加到目标加液孔对应的目标反应杯中。
这里，步骤S110和步骤S120为试剂添加过程。步骤S130和步骤S140为试剂采集过程。通常情况下，是先采集试剂再添加试剂，因此步骤S120要先于步骤S140。
在生化检测设备的整个实验检测过程中，均是试剂采集在先，试剂添加在后。在第一次采集时，其触发试剂采集的信号仅仅是目标试剂和/或目标样本转动至目标采样孔的信号。之后再采集时，其触发试剂采集的信号不仅仅是目标试剂和/或目标样本转动至目标采样孔的信号，还包括试剂添加完成状 态信号。
图11是本发明实施例十一的硫氧还蛋白还原酶(TR)检测的生化检测设备的操作方法流程示意图。
如图11所示，在实施例十的基础上，本发明实施例十一提供的用于硫氧还蛋白还原酶(TR)检测的生化检测设备的操作方法，还包括：
步骤S150，当状态感应装置检测到采样装置的试剂状态信息为试剂添加完成时，生成搅拌指令；
步骤S160，驱动装置基于接收到的搅拌指令，控制搅拌装置转动到目标加液孔以对目标反应杯中的混合液进行搅拌。
图12是本发明实施例十二的用于硫氧还蛋白还原酶(TR)检测的生化检测设备的操作方法流程示意图。
如图12所示，在实施例十一的基础上，本发明实施例十一提供的用于硫氧还蛋白还原酶(TR)检测的生化检测设备的操作方法，还包括：
步骤S170，当状态感应装置检测到搅拌装置的搅拌状态信息为搅拌完成时，生成搅拌复位指令；
步骤S180，驱动装置基于接收到的搅拌复位指令，控制搅拌装置复位。
图13是本发明实施例十三的用于硫氧还蛋白还原酶(TR)检测的生化检测设备的操作方法流程示意图。
如图13所示，本发明实施例十三提供的用于硫氧还蛋白还原酶(TR)检测的生化检测设备的操作方法，还包括：
步骤S210，当状态感应装置检测到反应装置转动预定角度的次数为预定次数时，生成试剂转动指令；
步骤S220，驱动装置基于接收到的试剂转动指令，控制容纳装置转动预定角度，使得下一个目标反应杯对准目标加液孔。
在本发明实施例十四提供的用于硫氧还蛋白还原酶(TR)检测的生化检测设备的操作方法中，还包括：
当驱动装置接收到用户通过用户操作指令输入界面输出的清洗指令时，基于接收到的所述清洗指令，控制所述清洗装置对目标加液孔对应的目标反应杯进行清洗。
下面结合人血中硫氧还蛋白还原酶活性检测的检测过程说明本发明的工作原理：
在本发明中，反应盘上反应杯的数量为81个，分为对照组和实验组。有两种分配方式，第一种情况：对照组的反应杯的数量为40个，实验组的反应杯的数量则为41个；第二种情况：对照组的反应杯的数量为41个，实验组的反应杯的数量为40个。81个反应杯分为9个组，每组反应杯包括9个反应杯。
容纳盘上具有三圈容纳固定件，每圈容纳固定件数量为40个。其中，第一圈容纳固定件用于盛放第一试剂瓶、第二圈容纳固定件用于盛放样本瓶，第三圈容纳固定件用于盛放第二试剂瓶。
本发明的最终目的是将81个反应杯中均加入第一试剂、样本和第二试剂。需要说明的是，本发明实施例中所涉及到的第一试剂、第二试剂和样本均为和硫氧还蛋白还原酶活性检测相关的试剂。其中，样本为血液、体液或组织匀浆液。第一试剂包括工作液(TrisHCL、吗啉基丙磺酸、磷酸氢二钠柠檬酸缓冲体系和磷酸氢二钠磷酸二氢钾缓冲溶液的混合溶液)和硫氧还蛋白 还原酶抑制剂的化合物。第二试剂为混合试剂，包括5,5-二硫代双(2-硝基苯甲酸)或取代6,6'-二硝基-3,3'-二硫代苯甲酸与烟酰胺腺嘌呤二核苷酸磷酸的混合溶液。
其中，工作液配置过程为：按照1:1:2:4的比例取试剂三羟甲基氨基甲烷盐酸盐TrisHCL(0.025-0.125mol/L,PH 5.5.8-7.2)，吗啉基丙磺酸(0.25mol/L)，磷酸氢二钠柠檬酸缓冲体系0.2mol/L，和磷酸氢二钠磷酸二氢钾缓冲溶液1-15mol/L；其中，所述磷酸氢二钠柠檬酸缓冲体系的PH为2.2-8.0；所述磷酸氢二钠磷酸二氢钾缓冲溶液的PH为4.9-8.2；然后将所述TrisHCL(三羟甲基氨基甲烷盐酸盐)、吗啉基丙磺酸、磷酸氢二钠柠檬酸缓冲体系和磷酸氢二钠磷酸二氢钾缓冲溶液混合均匀。其中，以1:1-1:5的比例混合所述工作液和抑制剂，形成所述抑制剂溶液；将所述抑制剂溶液混合均匀；其中所述抑制剂为硫氧还蛋白还原酶抑制剂化合物。可以为化学单体硒啉类化合物。以1:2-1:8的比例混合所述试剂A和所述试剂B，形成所述混合试剂；具体的，试剂A和试剂B的混合比例范围为1:2-1:8，优选的以1:4-5的比例来混合试剂A和试剂B，此时混合生成的混合试剂在检测时的准确度较高，其中，所述A试剂为5,5-二硫代双(2-硝基苯甲酸)或取代6,6'-二硝基-3,3'-二硫代苯甲酸；B试剂为烟酰胺腺嘌呤二核苷酸磷酸。该部分可参见同一申请人的另一件发明专利申请“一种人外周血硫氧还蛋白还原酶活性检测方法”。
关于采液孔的设置数量，当仅需要采集一种试剂时，则只需要在容纳盘盖上设置一个采液孔；当需要采集多种试剂时，则需要在容纳盘盖设置多个采液孔，使得每种试剂对应一个加液孔，这样设置的目的是加快采液速度。 在本发明的人血中硫氧还蛋白还原酶活性检测实验中，由于需要采集三种试剂，因此，本发明设置了三个采液孔，第一试剂对应第一采液孔，样本对应第二采液孔，第二试剂对应第三采液孔。
关于加液孔的设置数量，本申请为了提高加液的速度，故设置了两个加液孔(第一加液孔和第二加液孔)。在第一周期组时，只给第一组反应杯加第一试剂，可以通过第一加液孔或第二加液孔添加第一试剂。第二周期组时，给第一组反应杯加样本，给第二组反应杯加第一试剂，可以通过第一加液孔和第二加液孔添加第一试剂和样本。第三周期组时，给第一组反应杯加第二试剂，给第二组反应杯加样本，给第三组反应杯加第一试剂，之后，按照第三组的加液顺序，继续给第二组反应杯至第9组反应杯加入第一试剂、样本和第二试剂。
下面结合图14说明本发明检测硫氧还蛋白还原酶的实验过程：
S1，第一周期组的第一周期开始，驱动装置控制容纳盘转动，当状态检测装置50检测到容纳装置中第一试剂转动至第一加液孔或第二加液孔时且试剂状态信息为试剂添加完成状态时，生成第一试剂采集指令；
S2，驱动装置基于接收到的第一试剂采集指令，控制采样装置转动至盛放装置的第一采样孔采集第一试剂；
S3，当状态检测装置检测到第一组反应杯中的第一个反应杯转动至第一加液孔或第二加液孔时且状态感应装置50检测到采样装置30的试剂状态信息为试剂采集完成时，生成第一试剂添加指令；
S4，驱动装置70在接收到第一试剂添加指令后，控制采样装置30转动至反应装置20的第一加液孔或第二加液孔并将第一试剂添加到第一加液孔 或第二加液孔对应的第一个反应杯中。
S5，第一组反应杯中的第一个反应杯添加完第一试剂后，将第一组反应杯中的第二个反应杯转动至第一加液孔或第二加液孔，继续执行上述采集第一试剂和添加第一试剂的过程，直到将第一组反应杯中的9个反应杯全部添加完第一试剂，第一周期组结束。
S6，第二周期组的第一周期开始，驱动装置控制反应装置20转动，当状态感应装置50检测到反应装置中第二组反应杯中的第一个反应杯转动至第一加液孔时采样装置的试剂状态信息为试剂采集完成状态时，生成第一试剂添加指令。
S7，驱动装置基于接收到的试剂添加指令，控制采样装置转动至反应装置的第一加液孔并将第一试剂添加到第一加液孔对应的第一反应杯中；
S8，当状态感应装置检测到反应装置中采样装置的试剂状态信息为添加完成状态时，生成第一试剂采集指令；
S9，驱动装置基于接收到的第一试剂采集指令，控制采样装置转动至容纳装置的第一采样孔采集第一试剂；
S10，当状态检测装置检测到第二组反应杯中的第一个反应杯转动至第一加液孔或第二加液孔时且状态感应装置50检测到采样装置30的试剂状态信息为试剂采集完成时，生成第一试剂添加指令；
S11，驱动装置70在接收到第一试剂添加指令后，控制采样装置30转动至反应装置20的第一加液孔并将第一试剂添加到第一加液孔对应的第一个反应杯中。
采样装置按照上述采集第一试剂的过程采集样本，按照第二周期组的第 一个周期添加第一试剂、第二个周期添加样本的顺序，直到第二组反应杯中的9个反应杯加入第一试剂，以及第一组反应杯中的9个反应杯加入样本，第二周期组结束。
在第三周期组的第一个周期，为第三组反应杯添加第一试剂；第二个周期，为第二组反应杯添加样本；第三个周期，为第一组反应杯添加第二试剂，直到将第三组反应杯中的9个反应杯加入第一试剂，第二组反应杯中的9个反应杯加入样本试剂，第一组反应杯中的9个反应杯加入第二试剂，第三周期组结束。
按照第三周期组的加液顺序，将第4-9组反应杯中均加入第一试剂、样本和第三试剂，直至整盘检测完成或停止检测。该部分可参见同一申请人的另一件发明专利申请“一种硫氧还蛋白还原酶活性的协同检测设备和方法”。
需要说明的是，待每一组反应杯都加入了第一试剂、样本和第二试剂之后，即开始发生反应，反应时间最长为22个周期(每个周期22.5s)，优选地，为20个周期，对该组反应杯来说，在这22个周期内，若该组反应杯转动至加液位置，则移液装置静置处于等待状态。待该组反应杯反应结束后，可通过客户端发送指令控制液压装置，进而控制清洗装置将反应杯中反应后的废液抽走，并对反应杯进行清洗，清洗完成后该组反应杯即可继续进行加入第一试剂、样本和第二试剂的操作。
例如：对于第一组反应杯来说，第三周期组结束后，第一组反应杯内的包括第一试剂、样本和第二试剂的混合液即开始发生反应，此时，在下一个周期内，若反应盘的第一组反应杯转动至加液位置，则移液装置静置处于等待状态。
由于本发明的生化检测设备不仅可以对生化反应进行检测，还可以对非生化反应进行检测，因此，只需要在客户端的用户输入界面输入所需要做的 生化反应参数或非生化反应参数，即能够根据具体的生化反应或非生化反应的检测过程进行自动调整。
本发明旨在提供一种硫氧还蛋白还原酶活性检测方法，通过对适用于上述介绍的一种硫氧还蛋白还原酶活性的协同检测设备的样本/试剂体积的设定，能够符合运行的工作液体选择的方法要求，本发明的方法在上述介绍的一种硫氧还蛋白还原酶活性的协同检测设备上使用时，包括加样操作方法、避光操作要求方法、试剂混合的操作要求方法等针对驱动系统的智能指令方法规定。其中的智能指令如协同检测设备运行中每一周期组中周期数以及每一周期时间的规定，针对运行过程要求等都是和实现人外周血TR功能检测方法衔接的检测方法学。本发明的方法是适用于上述介绍的一种硫氧还蛋白还原酶活性的协同检测设备实现TR酶学检测功能要求的方法，用于上述介绍的一种硫氧还蛋白还原酶活性的协同检测设备的硫氧还蛋白还原酶活性检测方法。
本发明的另一目的还旨在保护一种人血中用于硫氧还蛋白还原酶生化检测的生化检测设备及其操作方法，一方面，本发明的生化检测设备首次实现了人血中硫氧还蛋白还原酶活性检测的全自动化检测，解决了现有技术中智能通过手工完成硫氧还蛋白还原酶活性检测的问题；另一方面，本申请的生化检测设备不仅能够实现单个样本整套TR检测流程，还能够实现多个样本连续完成TR检测流程。
具体地，1)通过硬件设置安排，包括反应盘的配置，容纳盘的配置，两盘之间的联动方式，加样和采样装置等，可以实现对于单一样本的整体检测流程的完成。由于对于样本的连续处理，连续加样和反应时间要求严格，由 于样本盘每转动两格，反应盘转动一格，即每个样本管取2次样本，分别加入两个反应杯，作为实验组和对照组数据的检测，因此能够完成单一样本的同步检测。
2)通过硬件设置安排和方法的改进，其中，硬件设置包括反应盘的配置，样本盘的配置，两盘之间的联动方式，加样和采样装置等，可以实现多样本连续检测流程的完成。方法的改进包括对于样本盘和反应盘所规定的转动时间，转动间隔，转动距离，采样顺序，加样时间等时序安排，由于每一组反应杯需要加三次溶液，因而可以在每一组反应杯加溶液的过程中同时开始下一组反应杯的加液，这样，能够使得多个样本连续进行检测，且可以循环检测，从而缩短检测时间。
此外，本发明采用特定驱动硬件、使得软件和硬件能够协同工作，可以提高临床TR检测效率、节省成本。且通过本发明的生化检测设备检测出的实验结果，能够满足国家检测标准要求。
应当理解的是，本发明的上述具体实施方式仅仅用于示例性说明或解释本发明的原理，而不构成对本发明的限制。因此，在不偏离本发明的精神和范围的情况下所做的任何修改、等同替换、改进等，均应包含在本发明的保护范围之内。此外，本发明所附权利要求旨在涵盖落入所附权利要求范围和边界、或者这种范围和边界的等同形式内的全部变化和修改例。

Claims (29)

  1. 一种硫氧还蛋白还原酶活性检测方法，其特征在于，包括：
    配液，
    配置工作液、抑制剂溶液和混合试剂；
    加样本，
    将50uL-70uL的所述工作液加入对照组反应杯；
    将50uL-70uL的所述抑制剂溶液加入实验组反应杯；
    向对照组反应杯和实验组反应杯分别加入10uL-30uL的样本；
    温育，
    在避光环境下，将所述对照组反应杯和所述实验组反应杯放置在30℃-40℃温度下孵育第一预定时间；
    测定，
    分别向所述对照组反应杯和所述实验组反应杯加入100uL-150uL的混合试剂；
    在预定波长下，测定第二预定时间段内的吸光度值。
  2. 根据权利要求1所述的检测方法，其特征在于，所述配置工作液的步骤包括：
    按照1:1:2:4的比例取三羟甲基氨基甲烷盐酸盐、吗啉基丙磺酸、磷酸氢二钠柠檬酸缓冲体系和磷酸氢二钠磷酸二氢钾缓冲溶液；
    将所述三羟甲基氨基甲烷盐酸盐、吗啉基丙磺酸、磷酸氢二钠柠檬酸缓冲体系和磷酸氢二钠磷酸二氢钾缓冲溶液混合均匀。
  3. 根据权利要求2所述的检测方法，其特征在于，
    所述三羟甲基氨基甲烷盐酸盐的PH为5.5-7.2，浓度为0.025-0.125mol/L；
    所述吗啉基丙磺酸的浓度为0.25mol/L；
    所述磷酸氢二钠柠檬酸缓冲体系的PH为2.2-8.0，浓度为0.2mol/L；所述磷酸氢二钠磷酸二氢钾缓冲溶液的PH为4.9-8.2，浓度为1-15mol/L。
  4. 根据权利要求1所述的检测方法，其特征在于，所述配置抑制剂溶液的步骤包括：
    以1:1-1:5的比例混合所述工作液和抑制剂，形成所述抑制剂溶液；
    将所述抑制剂溶液混合均匀；
    其中，所述抑制剂为硫氧还蛋白还原酶抑制剂化合物。
  5. 根据权利要求1-4中任一项所述的检测方法，其特征在于，所述配置混合试剂的步骤包括：
    以1:4-1:8的比例混合所述试剂A和所述试剂B，形成所述混合试剂；
    将所述混合试剂混合均匀；
    所述A试剂为5,5-二硫代双(2-硝基苯甲酸)或取代6,6'-二硝基-3,3'-二硫代苯甲酸；
    所述B试剂为烟酰胺腺嘌呤二核苷酸磷酸。
  6. 根据权利要求1所述的检测方法，其特征在于，所述预定温度为30-40℃。
  7. 根据权利要求1所述的检测方法，其特征在于，所述第一预定时间为8-20分钟。
  8. 根据权利要求7所述的检测方法，其特征在于，所述第一预定时间为10分钟。
  9. 根据权利要求1所述的检测方法，其特征在于，所述预定波长为405nm-450nm。
  10. 根据权利要求1或9所述的检测方法，其特征在于，所述第二预定时间为20-30个周期。
  11. 一种用于硫氧还蛋白还原酶活性检测设备，其特征在于，包括：
    容纳装置(10)，用于盛放多个试剂和/或样本，并在驱动装置(70)的驱动作用下，围绕轴心周期性转动使得目标试剂和/或目标样本转动至目标加液孔；
    反应装置(20)，用于盛放多个反应杯，并在驱动装置(70)的驱动作用下，围绕轴心周期性转动使得目标反应杯转动至目标采样孔；
    采样装置(30)，用于在驱动装置(70)的驱动作用下，围绕轴心周期性转动，将从目标采样孔中采集的所述目标试剂和/或目标样本加入目标加液孔对应的所述目标反应杯中；状态感应装置(50)，用于检测所述反应装置(20)的反应转动状态信息、容纳装置(10)的容纳转动状态信息、采样装置(30)的试剂状态信息；
    主控系统(60)，分别与所述状态感应装置(50)、驱动装置(70)连接，用于基于反应转动状态信息、容纳转动状态信息和试剂状态信息，生成并向驱动装置(70)发送相应的控制指令；
    驱动装置(70)，分别与所述采样装置(30)、所述容纳装置(10)、所述反应装置(20)连接，用于基于接收到的控制指令，控制所述采样装 置(30)、所述反应装置(20)、所述容纳装置(10)执行相应的操作。
  12. 根据权利要求11所述的检测设备，其特征在于，所述容纳装置(10)包括：
    容纳盘(11)，活动设置，围绕轴心做周期性转动；
    多个容纳固定件(12)，用于盛放试剂瓶和/或样本瓶。
  13. 根据权利要求1所述的生化检测设备，其特征在于，所述反应装置(20)包括：
    反应盘(21)，活动设置，围绕轴心做周期性转动；
    多个反应杯固定件(22)，用于盛放反应杯。
  14. 根据权利要求11所述的检测设备，其特征在于，所述采样装置(30)包括：
    采样旋转件(31)，活动设置，用于在驱动装置(70)的驱动作用下，围绕轴心周期性转动；
    采样固定件(32)，固定设置在采样转动件(31)上，在采样旋转件(32)的带动作用下转动；
    采样针(33)，一端为固定端，另一端为自由端。
  15. 根据权利要求11所述的检测设备，其特征在于，
    所述主控系统(60)在接收到目标反应杯转动至目标加液孔时的反应转动状态信息且试剂状态信息为试剂采集完成状态时，生成试剂添加指令；
    所述驱动装置(70)在接收到试剂添加指令后，控制采样装置(30)转动至反应装置(20)的目标加液孔并将目标试剂和/或目标样本添加到 目标加液孔对应的目标反应杯中。
  16. 根据权利要求11所述的检测设备，其特征在于，
    所述主控系统(60)在接收到目标试剂和/或目标样本是否转动至目标采样孔时的容纳转动状态信息时和试剂状态信息为试剂添加完成状态时，生成试剂采集指令；
    所述驱动装置(70)在接收到试剂采集指令后，控制采样装置(30)转动至盛放装置(10)的目标采样孔处采集目标试剂和/或目标样本。
  17. 根据权利要求11所述的检测设备，其特征在于，所述生化检测设备还包括：
    搅拌装置(40)，用于在驱动装置(70)的驱动作用下，围绕轴心做周期性转动以对目标反应杯中形成的混合液进行搅拌。
  18. 根据权利要求17所述的生化检测设备，其特征在于，所述搅拌装置(40)包括：
    搅拌旋转件(41)，活动设置，用于在驱动装置(70)的驱动作用下，围绕轴心周期性转动；
    搅拌固定件(42)，固定设置在搅拌转动件(31)上，在搅拌旋转件(42)的带动作用下转动；
    搅拌针(43)，一端为固定端，另一端为自由端。
  19. 根据权利要求17所述的检测设备，其特征在于，
    所述主控系统(60)在接收到试剂状态信息为试剂添加完成时，生成搅拌控制指令；
    驱动装置(70)在接收到搅拌控制指令后，控制搅拌装置(40)转动 到目标加液孔以对目标反应杯中的混合液进行搅拌。
  20. 根据权利要求17-19中任一项所述的检测设备，其特征在于，
    所述状态感应装置(50)还用于检测搅拌装置(40)的搅拌状态信息；
    所述主控系统(60)，用于在接收到搅拌状态信息为搅拌完成后，生成搅拌复位指令；
    驱动装置，与搅拌装置(40)连接，控制搅拌装置(40)复位。
  21. 根据权利要求11所述的检测设备，其特征在于，
    所述主控系统(60)在接收到转动预定角度的次数为预定次数时的反应转动状态信息时，生成试剂转动指令；
    容纳装置(10)在接收到试剂转动指令后，控制容纳装置(10)转动预定角度，使得下一个目标反应杯对准目标加液孔。
  22. 根据权利要求11所述的检测设备，其特征在于，还包括：
    清洗装置(90)，用于将清洗液中的液体输入所述目标加液孔对应的目标反应杯以对所述目标反应杯进行清洗。
  23. 根据权利要求22所述的检测设备，其特征在于，所述清洗装置(90)包括：
    液压装置(91)，用于控制清洗液中液体进入目标反应杯或将目标反应杯中的废液排出目标反应杯；
    清洗组件(92)，与所述液压装置(91)连接，在所述液压装置(91)的作用下，对目标反应杯进行清洗。
  24. 根据权利要求22或23所述的检测设备，其特征在于，还包括：
    客户端(110)，与所述主控系统(60)连接，用于提供用户操作指令 输入界面，并用于采集用户在所述用户操作指令输入界面输入的清洗指令，将所述清洗指令发送给所述主控系统(60)；
    所述驱动装置(70)在接收到所述清洗指令后，控制所述清洗装置(90)对目标加液孔对应的目标反应杯进行清洗。
  25. 一种硫氧还蛋白还原酶活性检测设备的操作方法，其特征在于，包括：
    当状态感应装置检测到反应装置中目标反应杯转动至目标加液孔且采样装置的试剂状态信息为试剂采集完成状态时，生成试剂添加指令；
    驱动装置基于接收到的试剂添加指令，控制采样装置转动至反应装置的目标加液孔并将目标试剂和/或目标样本添加到目标加液孔对应的目标反应杯中；
    当状态感应装置检测到容纳装置中目标试剂和/或目标样本转动至目标采样孔时且试剂状态信息为试剂添加完成状态时，生成试剂采集指令；
    驱动装置基于接收到的试剂采集指令，控制采样装置转动至盛放装置的目标采样孔处采集目标试剂和/或目标样本。
  26. 根据权利要求25所述的操作方法，其特征在于，还包括：
    当状态感应装置检测到采样装置的试剂状态信息为试剂添加完成时，生成搅拌指令；
    驱动装置基于接收到的搅拌指令，控制搅拌装置转动到目标加液孔以对目标反应杯中的混合液进行搅拌。
  27. 根据权利要求26所述的操作方法，其特征在于，还包括：
    当状态感应装置检测到搅拌装置的搅拌状态信息为搅拌完成时，生成 搅拌复位指令；
    驱动装置基于接收到的搅拌复位指令，控制搅拌装置复位。
  28. 根据权利要求26-27中任一项所述的操作方法，其特征在于，还包括：
    当状态感应装置检测到反应装置转动预定角度的次数为预定次数时，生成试剂转动指令；
    驱动装置基于接收到的试剂转动指令，控制容纳装置转动预定角度，使得下一个目标反应杯对准目标加液孔。
  29. 根据权利要求26-27中任一项所述的操作方法，其特征在于，还包括：
    当驱动装置接收到用户通过用户操作指令输入界面输出的清洗指令时，基于接收到的所述清洗指令，控制所述清洗装置对目标加液孔对应的目标反应杯进行清洗。
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