CN108120755B - Detection device and application thereof - Google Patents
Detection device and application thereof Download PDFInfo
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- CN108120755B CN108120755B CN201711385284.0A CN201711385284A CN108120755B CN 108120755 B CN108120755 B CN 108120755B CN 201711385284 A CN201711385284 A CN 201711385284A CN 108120755 B CN108120755 B CN 108120755B
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Abstract
The invention relates to the field of in-vitro diagnosis medical instruments, in particular to a detection device and a preparation method and application thereof. The invention discloses a POCT (point of care testing) diagnostic card. The calibration reagent, the reagent valve, the electrode array chip, the waste liquid storage cavity and the like are integrated on the diagnostic card, so that reagent sample adding is not needed any more in the detection process, extra medical wastes are not generated any more, all fluids and pipelines are integrated on the test card, the use convenience is improved to a great extent, and the blocking risk of an instrument liquid path is avoided. The diagnosis card applies the electrochemical principle, can be used for in vitro diagnosis of disease patients, analysis and detection of some biological and chemical substances in human blood samples, and can be applied to a plurality of medical diagnosis fields, such as biochemical item tests, blood and gas item tests, metabolite tests, hematology tests, blood coagulation tests, and immunology tests (cardiac markers and the like).
Description
Technical Field
The invention relates to the field of in-vitro diagnosis medical instruments, in particular to a detection device and application thereof.
Background
The POCT rapid diagnosis technology has become a development trend of medical diagnosis, and a plurality of corresponding diagnosis cards or diagnosis test strips are provided; mainly comprises colloidal gold test paper, immunofluorescence test paper, blood glucose test paper and the like, and the common characteristic of the test paper is disposable.
The colloidal gold test paper and the immunofluorescence test paper have low measurement precision due to the limitation of the test methodology adopted by instruments, and generally are qualitative or semi-quantitative measurements.
The electrochemical technology can realize the response, transmission and acquisition of signals through a pure hardware circuit, and along with the rapid development of the PCB manufacturing technology, the size of a multi-channel electrochemical tester can be made into the size of 1 palm, so that the multi-channel electrochemical tester is very suitable for the development requirement of the POCT rapid diagnosis technology; with the development of solid-state dry-type electrochemical sensor technology, highly integrated micro-electrochemical sensor arrays have been applied to clinical tests, such as blood glucose test, blood gas electrolyte test, and the like.
The blood glucose test paper is manufactured by printing a circuit layer on a PET substrate by adopting a screen printing technology, coating a chemical reagent on a sensor area formed by printing, and then drying and forming. The blood glucose test paper adopts an electrochemical principle and has the advantages of higher precision and small instrument; however, the blood glucose test paper is a single test item, and a plurality of test items cannot be completed on the same test paper strip; and the blood glucose test paper is not calibrated by a calibration solution before testing, so that the testing precision is limited.
Therefore, the detection device which is highly integrated, simple in process and convenient to operate has important practical significance.
Disclosure of Invention
In view of the above, the present invention provides a detection device and an application thereof. The detection device integrates a calibration reagent, an electrode sensor array, and all flow channels and control elements which are in contact with liquid, can be used for in-vitro diagnosis of disease patients based on an electrochemical principle, analyzes and detects some biological and chemical substances in human blood samples, and can be applied to a plurality of medical diagnosis fields, such as biochemical item tests, blood and gas item tests, metabolite tests, hematology tests, blood coagulation tests, and immunological tests (heart markers and the like).
In order to achieve the above object, the present invention provides the following technical solutions:
the invention provides a detection device, which comprises a substrate, wherein the substrate is integrated with a liquid micro-channel and an electrode sensor array, and the liquid micro-channel comprises a calibration reagent micro-channel, a sample micro-channel and an electrode sensor area micro-channel; the reagent storage component is tightly connected with the calibration reagent flow channel.
In some embodiments of the invention, the reagent storage component comprises at least two layers of film; the first layer of film is tightly arranged on the inner wall of the calibration reagent flow passage and is formed into a cavity through punch forming; and the second layer of film is arranged on the first layer of film in parallel, and is combined with the first layer of film through hot pressing or ultrasonic welding to form a completely sealed cavity, so that the reagent storage component is obtained.
In some embodiments of the invention, the electrode sensor array comprises a metal layer and an insulating layer;
the metal layer comprises electrochemically active electrode sensor sites and conductive contacts for electrode sensor signals;
the insulating layer is provided with through holes in the regions corresponding to the electrode sensor sites, well-shaped blind holes are formed in the insulating layer and the electrode sensor sites of the metal layer, and the electrode sensor array applicable to multi-parameter testing is obtained by coating selective sensor films on the well-shaped blind holes.
In some embodiments of the present invention, a valve plate is disposed at a connection position of the reagent storage part and the calibration reagent flow channel, and communication and separation between the reagent storage part and the calibration reagent flow channel are controlled by the valve plate.
In some embodiments of the invention, the test device further comprises a calibration reagent buffer member disposed between the reagent storage member and the calibration reagent flow path.
In some embodiments of the invention, the detection device further comprises a sample inlet and a microfluidic sample flow channel, and the distal blood sampling is performed by capillary siphon action or the sample is injected by a syringe.
In some embodiments of the present invention, a groove for removing air bubbles in the sample is disposed around the sample inlet of the detection device.
In some embodiments of the present invention, the sample inlet of the detection device is further provided with a C-shaped component, and the C-shaped component is tightly connected with the sample inlet, so as to reduce the resistance of the syringe to sample injection.
In some embodiments of the invention, the detection device further comprises a waste liquid recovery part, one end of the waste liquid recovery part is connected with the liquid micro-channel, and the other end of the waste liquid recovery part is communicated with the outside, so as to ensure smooth liquid flow.
The invention also provides application of the detection device in biochemical project testing, blood and gas project testing, metabolite testing, hematology testing, blood coagulation testing and immunology testing.
The invention discloses a POCT (point of care testing) diagnostic card. The calibration reagent, the reagent valve, the electrode array chip, the waste liquid storage cavity and the like are integrated on the diagnostic card, so that reagent sample adding is not needed any more in the detection process, extra medical wastes are not generated any more, all fluids and pipelines are integrated on the test card, the use convenience is improved to a great extent, and the blocking risk of an instrument liquid path is avoided. The diagnosis card applies the electrochemical principle, can be used for in vitro diagnosis of disease patients, analysis and detection of some biological and chemical substances in human blood samples, and can be applied to a plurality of medical diagnosis fields, such as biochemical item tests, blood and gas item tests, metabolite tests, hematology tests, blood coagulation tests, and immunology tests (cardiac markers and the like).
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below.
FIG. 1 shows a front projection view of a diagnostic card;
FIG. 2A shows a front projection view of a targeting agent zone;
FIG. 2B is a schematic cross-sectional view of a flow path of the targeting reagent zone;
FIG. 3 shows a front projection view of a sample inlet;
FIG. 4A is a schematic front projection view of a chip and a liquid channel;
FIG. 4B is a schematic cross-sectional view of the chip and the liquid channel;
FIG. 5A shows a response curve for a pH electrode;
FIG. 5B shows pO2A response curve of the electrode;
FIG. 6A shows comparative results of pH testing;
FIG. 6B shows pO2Testing the comparison result;
wherein, 1-sample inlet (sample injection port); 2-scaling the storage means (scaling reagent pack); 3-electrode sensor array (electrode sensor array chip area); 4-a valve; 5-waste liquid recovery device (waste liquid chamber); 6-a calibration reagent buffer section (reagent buffer zone); 7-a circular hole; 8-flow channel of reagent flow to sensor chip area; 9. 10-respectively reserving the calibrating reagents for filling at the head end and the tail end during heat sealing, and sealing the channels after filling; 11-round hole for removing air bubble between the aluminum plastic film and the diagnosis card substrate when pasting the aluminum plastic film; 12a and 12 b-an upper layer and a lower layer of aluminum plastic films with the same material; 13. 14-two layers of common plastic films; 15. 16-a bump; 17. 18-upper and lower card shells for contacting the instrument and the diagnostic card respectively; 19-C shaped small discs; 20. 23-electrode sensors of different shapes; 21-base material with good chip insulation performance; 22-sensor array chip area flow channels; 24-a gold plated metal layer; 25-27-three layers of insulating materials with concentric through holes, wherein the diameters of the open holes are increased from 25 to 27, and the open holes and the metal layers shown in the figure 24 are laminated together to form an inverted cone-shaped sensor area; 28-intramembrane reagent composition; 29-sensor membrane with electrochemical or biological activity; 30-double-sided adhesive tape for bonding the chip and the test card.
Detailed Description
The invention discloses a detection device and application thereof, and a person skilled in the art can appropriately improve process parameters by referring to the content. It is expressly intended that all such similar substitutes and modifications which would be obvious to one skilled in the art are deemed to be included in the invention. While the methods and applications of this invention have been described in terms of preferred embodiments, it will be apparent to those of ordinary skill in the art that variations and modifications in the methods and applications described herein, as well as other suitable variations and combinations, may be made to implement and use the techniques of this invention without departing from the spirit and scope of the invention.
In order to realize the diagnosis card with the structure and the functions, a diagnosis card base body with a liquid flow channel groove is designed firstly. The base material can be PET, PBT or PP material. The whole card can be divided into a calibration reagent area, an electrode sensor array area, a sample introduction area and a waste liquid storage area. In order to minimize the thickness of the whole test card and not to influence the smooth flow of liquid on the test card, the electrode sensor array area flow channels and other flow channels can be distributed on the front and back surfaces of the test card. The width of the flow channel of the sensor array area depends on the size of an electrode on the electrode sensor array chip, and is generally between 800 and 1500 mu m; the length and width of the flow channel of the calibration reagent zone determine the amount of reagent carried by the diagnostic card, and usually ranges from 75 to 110 μ L; the length and width of the sample introduction area flow channel and the sensor array area flow channel determine the size of the sample required by the test of the diagnostic card, and the sample size is generally 60-100 mu L; the design of the waste liquid area flow channel depends on the amount of the calibration reagent and the amount of the sample designed in the previous step, and in order to ensure that the calibration reagent or the sample does not overflow the test card in the test process, the capacity of the waste liquid cavity is designed to exceed 20% of the sum of the amount of the calibration reagent and the amount of the sample.
The main flow channel cavity of the calibration reagent area is designed into two parallel wide flow channels which are connected through a small flow channel. One end of the reagent valve is connected with the calibration reagent through a small flow channel, and the other end of the reagent valve is connected with the electrode sensor. In order to ensure the long-term stability of the reagent, especially the stability of O2 and CO2 gases contained in the blood gas calibration reagent, an aluminum plastic film is required to wrap the reagent cavity. The method adopts an adhesive mode to fix a layer of aluminum-plastic film on a diagnostic card base body, then forms an aluminum-plastic film/base body runner groove by punching, and also forms a groove for placing a valve plate in a valve plate area; then the second layer of aluminum-plastic film and the first layer of aluminum-plastic film are bonded together through heat sealing or ultrasonic welding to form a cavity with the periphery being closed, and the valve plate is also closed in the reagent bag; two channels are reserved at the head end and the tail end during heat sealing and used for filling the calibration reagent, and the channels are closed after filling is completed. The valve switch controls the flow of the calibration reagent to the electrode sensor area. Before use, the valve is in a normally closed state. After the diagnostic card starts the test, the valve is automatically opened through the test instrument, and then the two wide flow passage areas of the reagent area are pushed to enable the reagent to flow to the sensor array area.
The technical scheme adopted by the sensor array chip refers to the manufacturing technology of the smart card chip.
The technology of the smart card chip is quite mature, and due to the large demand of the electronic industry (such as a mobile phone SIM card chip and an IC card chip), the cost of the smart card chip is quite low.
The basic structure of a smart card chip includes an insulating layer and a metal layer. The insulating layer and the metal layer can be bonded by glue. The adhesive layer may be one or several layers.
Through holes are punched on the insulating layer and the adhesive layer, and then the sensor chip is bonded with the metal layer, so that various sensor chips with different structures can be formed. After bonding, the metal surface is exposed through the through-hole of the insulating layer to serve as a reaction contact surface of the bioelectrochemical sensor, and the material is generally Au or Pt. The well formed by the through hole of the insulating layer plays a key role in the preparation of the sensor film. Generally, the bioelectrochemical sensor is composed of multiple layers, and the thickness of each layer has different requirements, for example, the conductive electrolyte layer is generally several μm to several tens μm, the thickness of the ion selective layer is generally 20-40 μm, and the thickness of the air permeable layer is generally several μm. The well-shaped boundary formed by the through holes of the insulating layer or the bonding layer can enable the film formula solution to flow randomly during preparation of the sensor film, when the amount of liquid dripped into the boundary is constant every time, the thickness of the film is well controlled due to the fact that the area of the sensor formed by the boundary is limited to be constant, and the thickness of the film is a key parameter influencing the performance of the sensor. Due to the limitation of the boundary of the through hole, the strict surface plasma treatment on the contact surface of Au or Pt is not needed to achieve the aim of controlling the shape and the thickness of the film as in the conventional sensor film process. According to the requirements of specific sensors, insulating layers and adhesive layers with different thicknesses can be selected, and the size of the open through hole can also be designed according to actual requirements, so that a conductivity type sensor, a voltage type sensor and a current type sensor can be conveniently integrated on the same tiny chip (for example, 30mm multiplied by 10 mm).
The sensor layer and the signal acquisition connecting layer are respectively arranged on the front side and the back side of the same metal sheet, so that the size of the sensor chip is greatly reduced. The sensor layer is contacted with a sample to be tested, the size of the sensor can be designed into a micro size of hundreds of micrometers, a flow channel flowing through the surface of the sensor can also be designed into a width of hundreds of micrometers, and the sample size required by the test is very small (typical value: 20-100 mu L), so that a large amount of blood samples can be saved.
The chip is small and thin (the thickness is generally 0.15 mm-0.2 mm), so that the chip is very convenient for mass production. And optical detection marks can be printed on a chip screen for production batch identification, visual positioning, visual QC and the like during automatic production. The thickness of 0.15 mm-0.2 mm is also very convenient for subsequent cutting and sub-packaging of chips.
The microfluidic POCT diagnostic card provided by the invention can be used for in-vitro diagnosis of disease patients and analysis and detection of some biological and chemical substances in human blood samples by applying an electrochemical principle, and can be applied to a plurality of medical diagnosis fields, such as biochemical item tests, blood and gas item tests, metabolite tests, hematology tests, blood coagulation tests and immunological tests (cardiac markers and the like). Such a diagnostic card comprises:
a diagnostic card substrate: is a plastic card with the size and thickness equivalent to those of bank cards. The card is integrated with a liquid micro-channel, a calibration reagent storage area, a reagent valve, an electrode array mounting area and a waste liquid storage area.
Reagent bag: the diagnostic card is formed by pressing two aluminum-plastic films, wherein one aluminum-plastic film layer is tightly attached to the inner wall of a flow channel of a calibration reagent storage area on the diagnostic card, and the storage capacity of a reagent on the diagnostic card is determined by the size of a cavity formed by punch forming; the second layer of aluminum-plastic film is arranged on the first layer of aluminum-plastic film in parallel and is combined with the first layer of aluminum-plastic film through hot pressing or ultrasonic welding to form a completely sealed cavity.
Electrode array chip: and the lamination, printing and etching technologies of the flexible circuit board are adopted. Is formed by pressing a metal layer and a plurality of insulating layers; the metal layer is plated with gold on both sides, one side is an electrode sensor site with electrochemical activity, and the other side is a conducting contact of an electrode sensor signal. The insulating layer is provided with a through hole in the area corresponding to the electrode sensor site, and the through hole and the electrode sensor site on the metal layer form a well-shaped blind hole after being pressed. By applying various selective sensor membranes in the region of the well-shaped sensor sites, electrochemical sensors are obtained which can be used for measuring the corresponding parameters. A plurality of sensors are arranged in parallel to form a sensor array applicable to multi-parameter testing.
Preferably, the card has a size and a thickness equivalent to those of a bank card, and a calibration reagent pack, a reagent valve, an electrode sensor array, a calibration reagent microchannel, a sample microchannel, an electrode sensor zone microchannel, a waste liquid cavity and the like are integrated on the card.
Preferably, the sample and electrode sensor area micro-flow channel and the sample micro-flow channel are distributed on the upper and lower surfaces of the diagnostic card and are formed by sealing a groove on the substrate of the diagnostic card and a plastic film. The material of the diagnosis card substrate and the plastic film can be PET, PBT or PP.
Preferably, the calibration reagent bag consists of an upper aluminum plastic film and a lower aluminum plastic film; the calibration reagent pack ingeniously utilizes the cavity on the base material of the diagnostic card as a support, thereby not only ensuring the structural stability, but also being convenient for connecting the reagent pack with the test flow channel on the diagnostic card; in order to ensure the long-term stability of the reagent in the reagent pack, the minimum distance between the cavity of the reagent pack and the periphery of the test card is 5-10 mm.
Preferably, a valve plate is packaged at the joint of the reagent pack and the test flow channel, during testing, the sealed reagent pack is opened through the valve plate, the calibration reagent can flow to the electrode sensor array area, and the valve plate can be made of a stainless steel sheet with certain hardness or an epoxy resin sheet containing glass fibers.
Preferably, the diagnostic card further comprises a buffer for the calibration reagent. Opening the reagent bag by the valve of the reagent bag, and easily mixing the reagent with trace air in the valve area when the reagent begins to flow out, so that bubbles are mixed in the calibration reagent; the calibration reagent buffer of the diagnostic card is used for collecting the calibration reagent with air mixed at the front end.
Preferably, the diagnosis card is also integrated with an electrochemical sensor array, and each sensor site on the array corresponds to one measurement parameter, so that different clinical test item combinations can be realized based on different clinical requirements.
Preferably, the diagnostic card is also integrated with a microfluidic sample injection flow channel; the flow channel is subjected to corona or plasma surface treatment, and the inner surface of the flow channel is modified by hydrophilic polymer, so that the diagnosis card can sample peripheral blood through capillary siphon action. And simultaneously, the sample can be injected by a syringe.
Preferably, the injection port of the diagnostic card has a fine structure for reducing resistance during injection by the syringe and removing micro bubbles at the front end.
Preferably, the diagnosis card is also integrated with a waste liquid cavity; the tail end of the waste liquid cavity is communicated with the atmosphere, so that the smooth flowing of the liquid is ensured.
With reference to FIGS. 1-6A/6B, embodiments of the present invention and its use in vitro diagnostic procedures are described in detail.
Fig. 1 is a front projection view of a diagnostic card. FIG. 1 shows a sample injection port. FIG. 2 shows a calibration kit comprising several cavities of different lengths and widths. A round reagent valve (figure 4) at the top of the calibration kit; connecting the electrode sensor array chip area (figure 3) to a reagent buffer area (figure 6) through a reagent valve; the reagent buffer (FIG. 6) serves to exclude the calibration reagent and air bubbles from the initial extrusion, ensuring the reliability of the test results. FIG. 7 shows the intersection of the sample injection channel and the calibration reagent channel, the calibration reagent channel is on the upper layer, the sample injection channel is on the lower layer, and the sample enters the sensor testing area through the small round hole shown in FIG. 7. The diameter of the round hole in the figure 7 is 200-. After the test is completed, the discarded calibration solution and the sample are stored in the waste liquid chamber shown in FIG. 5. The upper right corner of the test card is provided with an oblique angle for foolproof design.
FIG. 2A is a front projection view of the targeting agent zone. Firstly, a layer of aluminum-plastic film is pasted on a diagnostic card substrate from bottom to top by adopting an adhesive mode, then an aluminum-plastic film/substrate runner groove (an area shown in a diagram 2) is formed by punching, and a circular hole shown in a diagram 11 is used for removing air bubbles between the aluminum-plastic film and the diagnostic card substrate when pasting the aluminum-plastic film, so that the aluminum-plastic film is guaranteed to be smoothly and flatly pasted on the inner wall of the groove; meanwhile, a groove is also formed in the circular area shown in the figure 4, the valve plate is placed in the groove, the second layer of aluminum-plastic film and the first layer of aluminum-plastic film are bonded together through heat sealing to form a cavity with the periphery being closed, and the valve plate is also closed in the reagent bag; two channels (shown in figures 9 and 10) are reserved at the head end and the tail end for filling the calibration reagent during heat sealing, and the channels are closed after filling. The flow channels shown in FIGS. 6 and 8 are the flow channels for the reagent buffer zone and the flow channels for the reagents to the sensor chip zone, respectively, located on the upper surface of the diagnostic card and connected to the calibration reagent flow channels located on the lower surface of the diagnostic card through the valves shown in FIG. 4.
FIG. 2B is a cross-sectional view along AA' of FIG. 2A. The figure 12a and the figure 12b are aluminum plastic films with the same material on the upper layer and the lower layer. The two layers of common plastic films, which can be made of PP or PET, are respectively adhered to the upper and lower surfaces of the diagnostic card, and form a closed liquid flow channel together with the grooves on the upper and lower surfaces of the diagnostic card. During testing, the diagnostic card is inserted into the instrument, and the upper and lower card cases where the instrument contacts the diagnostic card are shown in fig. 17 and fig. 18, respectively. Upon receipt of a test card insertion trigger signal, the instrument actuates the projections shown in FIG. 16 via the electrodes to move upwardly, thereby opening the reagent valve shown in FIG. 4 and allowing the reagent pack to communicate with the channel shown in FIG. 8. The tab shown in FIG. 15 is then driven upward by the electrodes, compressing the reagent pack area shown in FIG. 2, causing the calibration reagent to flow through channel 8 to the sensor area for calibration testing. Since the area shown in FIG. 4 usually has a small amount of air left, the test results are affected, especially by pO2、pCO2Testing of such gas items requires venting of the initial extruded calibrator reagent. This problem is best solved by the calibration reagent buffer shown in FIG. 2A, diagram 6. Since the channel width (800-.
Fig. 3 is a front projection view of the injection port. And a plurality of small grooves are formed around the sample inlet and used for removing a small amount of air bubbles possibly brought in the sample. The small C-shaped disk shown in figure 19 may serve to reduce drag during syringe injection.
The injection port flow channel is designed to be in a micron-scale, corona or plasma surface treatment is carried out on the flow channel, and hydrophilic polymer modification is carried out on the inner surface of the flow channel so as to ensure that the diagnostic card can carry out peripheral blood sampling through capillary siphon action.
FIG. 4A is a schematic front projection view of a chip and a liquid channel. The sensor array chip area channels are formed by attaching the chip to corresponding grooves on the diagnostic card, as shown in fig. 22, and the channels are typically 800 μm to 2mm wide. The illustrations 20 and 23 show different shapes of electrode sensors, one for each circular dot or elongated dot in fig. 4A, and a maximum of 14 different types of sensors can be integrated on the array chip shown in the present invention. Fig. 21 shows a base material with good chip insulation performance, which is generally an epoxy resin or polyimide material.
FIG. 4B is a cross-sectional view of the chip of FIG. 4A taken along direction BB'. The diagram 24 is a gold plated metal layer with the upper layer in contact with the sensor membrane to generate an electrical signal and the lower layer as a signal lead-out contact. The figures 25, 26 and 27 are three layers of insulating materials with concentric through holes, the diameters of the through holes are increased from 25 to 27, and the three layers of insulating materials are laminated with the metal layer shown in the figure 24 to form a sensor region with an inverted cone shape. The graphic 30 is a double-sided adhesive tape for bonding a chip and a test card, and the thickness is usually 0.05-0.08 mm. The diagram 29 shows a sensor membrane with electrochemical activity or biological activity, and the test items and performance of the sensor completely depend on the formulation, thickness, surface energy, uniformity, etc. of the membrane 29. The main component of the sensor membrane is usually some doped high molecular polymer; these membranes generally have good waterproof and air permeable properties, and can effectively prevent the reagent components (shown in figure 28) in the membrane from flowing outwards, and have good water vapor and pO2、pCO2Equal gas permeability; so as to ensure that the sensors can change from a dry state to a wet state when the liquid sample flows through the testing channel, thereby forming an electrochemical measuring loop with good conductivity.
The detection device and the raw materials and reagents used in the application of the detection device can be purchased from the market.
The invention is further illustrated by the following examples:
example 1 at pH and pO2The present invention will be described in detail by taking an electrode sensor as an example
preparing a pH electrode:
the pH electrode structure is as follows: an Ag/AgCl internal reference/electrolyte layer/ion-selective membrane layer;
the solid electrolyte adopted by the invention takes PVP or PVA as a solid phase carrier, 100-500mM NaCl or KCl is added as a supporting electrolyte, and PBS or Hepes is added as a pH buffer reagent;
the ion selective membrane is composed of: PVC (66% by wt.%) + NPOE (33% by wt.%) + H + support 1 (1% by wt.%)
And uniformly coating each sensor film layer on a circular area of the sensor chip in a micro dispersion manner.
pO2Preparing an electrode:
pO2the electrode structure is as follows: electrolyte membrane layer/gas permeable membrane layer;
the electrolyte layer uses hydrophilic gel as solid phase support carrier, such as PEG, trehalose, D4, etc., and KCl or NaCl is added as support electrolyte, and Hepes is added as pH buffer reagent;
the gas permeation film layer is preferably a polydimethylsiloxane-type polymer;
and uniformly coating each sensor film layer on a circular area of the sensor chip in a micro dispersion manner.
Preparing a calibration reagent:
50mM Hepes +150mM NaCl + preservative, adjusting pH to 7.35 with NaOH, and then passing a gas equilibrated with 21% O2+ N2 to obtain pO in the calibration reagent2Is 150 mmHg.
Electrode test curve
FIG. 5 shows integrated pH electrode and pO2Test response curves for the electrodes. After the diagnostic card is inserted into a test analyzer GSI-100 which is independently developed by the company, the analyzer automatically opens a reagent valve and pushes calibration solution to enter an electrode sensor area; when the sensor detects that the calibration solution completely covers the electrode area, data acquisition is started, namely a zero point on a test curve; at this point, sample preparation can begin, and the instrument monitors the change in sensor signal until after 60 seconds, after the slope of the curve has stabilized, the instrument prompt can begin to advanceAnd (5) sampling. The sample introduction time is required to be controlled within 60-240s, and if the sample introduction time exceeds 240s, the instrument will warn that the optimal sample introduction time is over, and please replace the diagnostic card. FIG. 5A shows an example of a response curve for a pH electrode with an electrode response sensitivity of 54-62mV/pH at 37 ℃; FIG. 5B is pO2Example of a response curve for an electrode. By testing pH and pO at different known concentration gradient values2Can establish a pH electrode and a pO separately2Scaling curves for the electrodes.
pH electrode scaling principle:
Δ V ═ s (pH) according to nernst equationSample(s)–pHScaling)，
So pH samples ═ Δ V × s '+ b'; by testing a series of known pH valuesi(i＝0,1,2……)Δ V at concentration samplei(i＝0,1,2……)Establishing a calibration curve to obtain a slope s 'and an intercept b';
when an unknown sample is tested, the pH value of the unknown sample can be obtained only by substituting the electrode response signal delta V.
pO2Electrode scaling principle:
defining a normalized scaling signal: d ═ iSample(s)/iScaling(i.e., the reaction current in the sample divided by the reaction current in the calibration solution), then:
pO2, sample of＝pO2, scaling*s*(D–a)/(1–a)
By testing a series of known pO2,i(i＝0,1,2……)D under the concentration samplei(i＝0,1,2……)Establishing a calibration curve to obtain a slope s and an intercept a;
when an unknown sample is tested, only the response signal D of the electrode is substituted, and the pO of the unknown sample can be obtained2The value is obtained.
pH and pO2Linear correlation results for electrode testing:
FIG. 6 is a graph showing pH and pO measurements using the GSI-100 test analyzer developed by the present company independently using the diagnostic card of the present invention2The results are compared with ABL800 test results of the industry post, Denmark Redmott Corp.
The results of the pH test shown in FIG. 6A andpO shown in FIG. 6B2The comparison of the test results shows that the correlation coefficient R2Are all larger than 0.99, and have good linear correlation.
The foregoing is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, various modifications and decorations can be made without departing from the principle of the present invention, and these modifications and decorations should also be regarded as the protection scope of the present invention.
Claims (4)
1. A detection device comprises a substrate, wherein the substrate is integrated with a liquid micro-channel and an electrode sensor array, and the liquid micro-channel comprises a calibration reagent micro-channel, a sample micro-channel and an electrode sensor area micro-channel; the device is characterized by also comprising a reagent storage component, wherein the reagent storage component is tightly connected with the calibration reagent micro-channel;
the reagent storage component comprises at least two layers of membranes; the first layer of membrane is tightly arranged on the inner wall of the calibration reagent micro-channel, and a cavity is formed by punch forming; the second layer of film is arranged on the first layer of film in parallel and is combined with the first layer of film through hot pressing or ultrasonic welding to form a completely sealed cavity, so that the reagent storage component is obtained;
the reagent buffer component is arranged between the reagent storage component and the calibration reagent micro-channel;
the electrode sensor array comprises a metal layer and an insulating layer;
the metal layer comprises electrochemically active electrode sensor sites and conductive contacts for electrode sensor signals;
the insulating layer is provided with through holes in the regions corresponding to the electrode sensor sites, well-shaped blind holes are formed in the insulating layer and the electrode sensor sites of the metal layer, and a selective sensor film is coated on the well-shaped blind holes to obtain an electrode sensor array applicable to multi-parameter testing;
a valve plate is arranged at the joint of the reagent storage part and the calibration reagent micro-channel, and the communication and the partition of the reagent storage part and the calibration reagent micro-channel are controlled by the valve plate;
the device also comprises a sample inlet and a microfluidic sample injection flow channel, and peripheral blood sampling is carried out through capillary siphon action or sample injection is carried out through an injector;
the sample inlet is still provided with C type part, C type part with the sample inlet is close connection, reduces the resistance that the syringe advances a kind.
2. The detection device according to claim 1, wherein a groove for removing air bubbles in the sample is arranged around the sample inlet.
3. The detecting device according to claim 1 or 2, further comprising a waste liquid recovering member, wherein one end of the waste liquid recovering member is connected to the liquid micro flow channel, and the other end is communicated with the outside, so as to ensure smooth liquid flow.
4. Use of the test device according to any one of claims 1 to 3 for biochemical tests, blood gas tests, metabolites tests, hematology tests, blood coagulation tests, immunological tests.
Priority Applications (1)
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| CN108918433B (en) * | 2018-07-26 | 2021-01-26 | 京东方科技集团股份有限公司 | Microfluid detection device |
| WO2020019809A1 (en) | 2018-07-26 | 2020-01-30 | Boe Technology Group Co., Ltd. | Microfluidic apparatus, method of detecting substance in microfluidic apparatus, and spectrometer |
| CN109444394B (en) * | 2018-10-29 | 2022-02-11 | 深圳市国赛生物技术有限公司 | Blood gas biochemical analyzer |
| CN109682960B (en) * | 2019-02-26 | 2023-09-29 | 苏州首通科技发展有限公司 | Test piece for electrolyte analyzer |
| CN113340952B (en) * | 2021-06-10 | 2024-01-30 | 南京晶捷生物科技有限公司 | Instant diagnosis testing device and application thereof |
| CN114354714B (en) * | 2022-01-07 | 2022-09-13 | 海南德名声科技有限责任公司 | Micro-fluidic chip of blood gas analyzer and application |
| CN115406946B (en) * | 2022-08-16 | 2023-06-30 | 山东卓越生物技术股份有限公司 | PO (potential of Hydrogen) of disposable instant detection card and sensor chip 2 Electrode preparation method |
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