KR101995253B1 - A device and a method for measuring a level of HbA1c of cartridge type - Google Patents

Abstract

The present invention relates to an apparatus and a method for detecting cartilage glycated hemoglobin (HbA1c), and more particularly, to a cartridge type glycated hemoglobin (HbA1c) detecting apparatus and method which is characterized in that a reaction liquid and a reaction substance necessary for the detection of glycated hemoglobin are independently implemented. The solution does not remain in the channel and the fluid is not mixed or overflowed, so that the content of hemoglobin and glycated hemoglobin can be more accurately measured. In other words, the present invention minimizes the effects of errors caused by driving each reaction liquid of the cartridge in a common channel and other interference substances in the blood, by individually controlling each fluid such as a reaction liquid and a sample There is an effect.

Description

Technical Field [0001] The present invention relates to a cartridge type glycated hemoglobin detecting device and a method thereof,

The present invention relates to an apparatus and a method for measuring glycated hemoglobin, and more particularly, to an apparatus and method for measuring glycated hemoglobin independently of a reaction liquid and a reaction substance necessary for detecting glycated hemoglobin in a cartridge, and more particularly, It is another object of the present invention to provide an apparatus and a method which can more accurately measure the hemoglobin and the content of glycated hemoglobin by preventing the solution from remaining in the channel and preventing the fluid from mixing or flooding.

Patients with type 1 or type 2 diabetes should routinely check for blood glucose (blood glucose) to prevent acute complications such as hypoglycemia, hypoglycemic shock from hyperglycemia, and hyperglycemic hemorrhagic acid. In addition, it is necessary to prevent the complications such as hemodialysis and artificial renal transplantation due to blindness due to diabetic retinopathy or diabetic nephropathy by preliminarily diagnosing or managing chronic vascular complications that may be caused by exposure to long-term hyperglycemia.

In recent years, the correlation between these complications and HbA1c and Glycated hemoglobin has been identified, and it has been used as an index of diabetes management by measuring glycated hemoglobin periodically. In addition, the American Diabetes Association (ADA) Is an indispensable indicator for the diagnosis of diabetes. Glycated hemoglobin refers to a form of irreversible binding of hemoglobin to the valine of the hemoglobin due to exposure to blood sugar for a long period of time, which is the protein that occupies the most part of the red blood cells of the blood. Since erythrocytes in the blood have a life cycle of 90 days to 120 days, glycated hemoglobin is an indicator of how much blood glucose is administered for 3 months.

In general, glycated hemoglobin is expressed as a percentile of glycated hemoglobin bound to valine in total hemoglobin. Therefore, the total hemoglobin concentration and the glycosylated hemoglobin concentration should be measured and calculated as percentiles.

Although the measurement of glycosylated hemoglobin is important, in the first medical institution, the frequency of measurement is insufficient to use a precise large-sized medical device, and a simple point-of-care testing device is required. However, since currently commercially available POCT-type devices emphasize the ease of use, the accuracy of measurement devices without hemoglobin separation is reduced. In addition, a device with a relatively high accuracy uses an expensive fluorescent material or Phenyle dye in the disposable cartridge to measure the glycated hemoglobin concentration in an optical manner, so that the cartridge becomes expensive, and accordingly, it is not suitable for use on the front line It is true.

In addition, in the conventional cartridge type glycated hemog- ene detecting apparatus, there is an inconvenience that a separate device is required to supply a part of reaction liquid, washer liquid, etc. from a pump outside the cartridge, , So that a continuous or dependent driving process is required between each reaction solution. That is, there is a difficulty in controlling the time and amount of the next reaction solution flowing after flowing a single reaction solution in the channel of the cartridge, or if the reaction solution or the like remains in the channel, And the amount of hemoglobin and the amount of glycated hemoglobin are greatly changed due to the influence of interference substances.

SUMMARY OF THE INVENTION The present invention has been made in order to solve the above-mentioned problems, and it is an object of the present invention to provide a method for measuring a hemoglobin concentration and a glycated hemoglobin concentration by more accurately measuring the content of hemoglobin and glycated hemoglobin by preventing a solution from remaining in a channel, The purpose is to do.

It is another object of the present invention to provide an apparatus and method for measuring HbA1c capable of minimizing an error caused by driving each reaction liquid in a cartridge in a common channel and the influence of other interference substances in blood.

According to an aspect of the present invention, there is provided a cartridge type glaucoma hemoglobin detecting apparatus comprising: a metal ion storage unit including a metal ion capable of binding to hemoglobin (Hb) contained in a sample; A membrane storage part including a membrane for separating the sample reacted with the metal ion into hemoglobin bound to the metal ion and a material not bound to the metal ion; A buffer solution reservoir containing a buffer solution for separating the hemoglobin bound to the metal ion into a metal ion and hemoglobin; A conversion material storage unit including a conversion material that converts the reacted hemoglobin into methylated hemoglobin (Met Hb) by reacting with the hemoglobin in which the metal ion is separated; Absorbance measurement means for measuring the absorbance of the hemoglobin of the methylated hemoglobin present in the converted substance storage section and measuring the hemoglobin level; An oxidase storage unit including an oxidase in which the metal ions are mixed with hemoglobin from which the metal ions are separated; An electrode unit in which a sugar solution capable of binding with the sugar of the hemoglobin and the oxidase is immobilized by reacting with a mixture of the hemoglobin and the oxidase in which the metal ions are separated; A signal generating material storage part including a signal generating material for generating an electric signal by reacting with hemoglobin and oxidizing enzyme bound to the electrode part; And an electrical signal measurement means for measuring an electrical signal generated at the electrode portion and measuring an amount of HbA1c.

Here, it is possible that the present invention further comprises a first waste solution storage part in which a surplus fluid of a sample containing the hemoglobin can be contained.

And a second waste solution storage part communicating with the membrane storage part, wherein the substance not bound to the metal ion is moved and stored.

The apparatus may further include a first washer fluid reservoir for washing the sample containing hemoglobin bound to the metal ion.

In addition, the conversion material contained in the conversion material storage unit is preferably solidified.

It is also possible to further include a second washer liquid reservoir for washing the sample containing the hemoglobin from which the metal ions are separated.

The third waste solution storage unit may further include a third waste solution storage unit communicating with the electrode unit, wherein the third waste solution storage unit moves and stores the substance not coupled to the sugar binding unit.

In addition, the present invention provides a method for producing a membrane-electrode assembly, which comprises at least one or more than one selected from the group consisting of the metal ion storage section, the membrane storage section, the buffer solution storage section, the conversion substance storage section, And a fluid transferring means for transferring the fluid.

In addition, the fluid transferring means may include a fluid loading unit for loading fluid and a syringe unit for loading the fluid.

Also, the fluid transfer means may include a position sensing unit for sensing a position corresponding to each of the metal ion storage unit, the membrane storage unit, the buffer solution storage unit, the conversion substance storage unit, the oxidizing enzyme storage unit, And may further comprise means.

The present invention further includes a control unit for automatically moving the fluid transfer unit according to the position sensing unit.

According to another aspect of the present invention, there is provided a method of preparing a sample, comprising: injecting a sample into a metal ion storage portion containing a metal ion capable of binding hemoglobin (hemoglobin, Hb); Injecting a sample reacted with the metal ion into a membrane storage section including a membrane separating the hemoglobin bound to the metal ion and the material not bound to the metal ion; Introducing the buffer solution from a buffer solution storage part containing a buffer solution for separating hemoglobin bound to metal ions into metal ions and hemoglobin and injecting the buffer solution into the membrane storage part; Injecting the hemoglobin into which the metal ions have been separated by the buffer solution into a conversion material storage section containing a conversion material for converting the hemoglobin into methylated hemoglobin (Met Hb); Measuring an absorbance of the hemoglobin of methylated hemoglobin present in the converted substance storage unit and measuring an amount of hemoglobin; Injecting hemoglobin into which the metal ions have been separated by the buffer solution into an oxidase storage unit containing the oxidase; Injecting a mixture of hemoglobin and oxidase in which the metal ions have been separated into an electrode unit in which sugar binding means capable of binding to the hemoglobin and oxidizing enzyme is immobilized; Injecting the signal generating material into the electrode unit from a signal generating material storage unit containing a signal generating material which reacts with hemoglobin and oxidizing enzyme to generate an electric signal; And an electrical signal measuring step of measuring an electrical signal generated from the electrode unit and measuring an amount of HbA1c (HbA1c).

The details of other embodiments are included in the detailed description and drawings.

The present invention is characterized in that the reaction solution and the reaction material necessary for the detection of glycated hemoglobin are independently implemented. Therefore, the solution does not remain in the channel and the fluid is not mixed or overflowed It is possible to more accurately measure the content of hemoglobin and the glycated hemoglobin.

In addition, the present invention can minimize the influence of errors generated by driving each reaction solution of the cartridge in a common channel and other interference substances in the blood, by individually controlling each fluid such as a reaction solution or a sample There is an effect.

BRIEF DESCRIPTION OF THE DRAWINGS Fig. 1 is a configuration diagram showing a configuration of a cartridge type glycated hemoglobin detecting apparatus according to a preferred embodiment of the present invention,
FIG. 2 is a schematic view for explaining an example of a state where metal ions are bound to hemoglobin according to the present invention,
FIG. 3 is a conceptual view for explaining an example of separation of hemoglobin bound with metal ions in the membrane according to the present invention, and FIG.
4 is a perspective view showing an example of the configuration of the fluid conveying means according to the present invention,
5 is a schematic view for explaining an example of a state in which the fluid transfer means according to the present invention is moved to each storage portion of the cartridge through the position sensing means,
FIG. 6 is a flowchart showing a process of a method for detecting glycated hemoglobin according to another preferred embodiment of the present invention,
FIG. 7 is a graph showing the results of hemoglobin content measurement by an optical method according to an embodiment of the present invention,
FIG. 8 is a graph showing the results of measuring the glycated hemoglobin content in an electrochemical manner according to an embodiment of the present invention.

BRIEF DESCRIPTION OF THE DRAWINGS The present invention is capable of various modifications and various embodiments, and specific embodiments are illustrated in the drawings and will be described in detail in the detailed description. It is to be understood, however, that the invention is not to be limited to the specific embodiments, but includes all modifications, equivalents, and alternatives falling within the spirit and scope of the invention. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS Hereinafter, the present invention will be described in detail with reference to the accompanying drawings.

The terminology used in this application is used only to describe a specific embodiment and is not intended to limit the invention. The singular expressions include plural expressions unless the context clearly dictates otherwise. In the present application, the terms "comprises" or "having" and the like are used to specify that there is a feature, a number, a step, an operation, an element, a component or a combination thereof described in the specification, But do not preclude the presence or addition of one or more other features, integers, steps, operations, elements, components, or combinations thereof.

The terms first, second, etc. may be used to describe various components, but the components should not be limited by the terms. The terms are used only for the purpose of distinguishing one component from another.

BRIEF DESCRIPTION OF DRAWINGS FIG. 1 is a configuration diagram showing a configuration of a cartridge type glycated hemoglobin detecting apparatus according to a preferred embodiment of the present invention. FIG.

The cartridge-type glycated hemoglobin detecting apparatus according to the present invention is for measuring the glycated hemoglobin concentration, and for this purpose, the total amount of hemoglobin contained in a sample such as blood or blood cells and the amount of glycated hemoglobin contained therein are measured separately. It is preferable to be a POCT-type miniaturization device having a microfluidic or strip-shaped cartridge structure.

The apparatus for detecting glycated hemoglobin according to the present invention basically comprises a metal ion storage part 10, a membrane storage part 20, a buffer solution storage part 30, a conversion substance storage part 40, An oxidizing enzyme storage unit 50, an electrode unit 60, a signal generating substance storage unit 70, and an electric signal measuring unit (not shown). Further, it may further include a waste solution storage unit 80 and a washer liquid storage unit 90, and it is possible to further include the fluid transfer unit 100.

Each of the above-described storage units may have a space or a groove or a hole shape contained in the cartridge or outside as shown in FIG. 1, and each of the storage units It is preferable that the reservoir portion is divided into separate independent spaces so as not to communicate with each other, and it is possible to have a hole or an injection port for allowing the fluid to flow in and out of the reservoir portion. Therefore, the movement of the reaction liquid and the like contained in each of the storage units can be performed manually or automatically by the separate fluid transfer means 100. The present invention is characterized in that the reaction solution and the reaction material necessary for the detection of glycated hemoglobin are independently embodied in the cartridge, and the concentration of glycated hemoglobin is more accurately measured through the process.

Each storage unit of the present invention will be described in detail.

First, the metal ion storage unit 10 includes a metal ion capable of binding hemoglobin (Hb) contained in a sample. That is, the metal ion may be a +2-valent metal ion capable of binding to porphyrin of hemoglobin (and glycosylated hemoglobin) as a means capable of binding with hemoglobin, and may be copper (Cu) or zinc (Zn) ion. These metal ions may be present in a solid state in the metal ion storage part 10 or temporarily immobilized therein, and more preferably in a liquid state for easy reaction with the sample. Therefore, when the sample flows into the metal ion storage part 10, the metal ions contained in the metal ion storage part 10 can bind to the hemoglobin in the sample.

Herein, FIG. 2 is a schematic view for explaining an example of a state where a metal ion is bound to hemoglobin according to the present invention, and more specifically, a chemical structural formula showing a state in which iron is removed from hemoglobin porphyrin. Hereinafter, the binding of metal ions to hemoglobin will be described in detail. In other words, porphyrin of hemoglobin essentially contains iron (Fe) atoms in the middle thereof, as shown on the left side of FIG. 2, but the present invention allows the metal ion to bind to the porphyrin instead of the iron (Fe) The hemoglobin can be bound by metal ions. For this, the metal ion storage unit 10 according to the present invention may further include a cleavage material for cleaving iron (Fe) atoms in porphyrin of hemoglobin, and the cleavage material may include glycine and magnesium chloride (MgCl ₂ ) . ≪ / RTI > Thus, the iron (Fe) atom is removed from the porphyrin by the cutting material, and the metal ion according to the present invention can be bonded to the Free Base Porphyrin from which the iron (Fe) atom has been removed.

The membrane storage part 20 includes a membrane for separating the sample reacted with the metal ion into hemoglobin bound to the metal ion and a substance not bound to the metal ion. That is, the membrane separates and collects only hemoglobin by metal ions, and functions to filter out other interfering substances (for example, plasma, protein, lipid, electrolyte, etc.) other than hemoglobin contained in the sample. In order to filter out the metal ions and the aggregated hemoglobin, the membrane should preferably be selected in terms of the material and pore size of the membrane, and the membrane (product of PALL as a super micro polysulfone material) It is more suitable to have the function of increasing the surface area of the base.

FIG. 3 is a conceptual view for explaining an example of separating hemoglobin in which the metal ions 11 are bound in the membrane 21 according to the present invention. As shown in FIG. 3, the membrane 21 has metal ions 11, And the saccharification protein such as glycated albumin not bound by the metal ion 11 and the saccharide and glucose are removed from the sample without passing through the hemoglobin bound by the metal ion 11, . The blood sample flowing into the metal ion storage part 10 includes hemoglobin (and / or hemoglobin bound to metal ions) as well as other interference substances. The sample is transferred to the membrane storage part 20 Here, only hemoglobin can be collected.

The buffer solution storage part 30 includes a buffer solution for separating the hemoglobin bound to the metal ions into metal ions and hemoglobin. That is, the buffer solution reacts with the hemoglobin to which the metal ion is bound to separate the metal ion from the hemoglobin, and among them, it is most effective to use phosphate buffered saline (PBS). The present invention seeks to further improve the accuracy by separating hemoglobin and metal ion through the buffer solution, thereby preventing errors in the hemoglobin amount measurement by the metal ions. The buffer solution may be transferred from the buffer solution storage part 30 to the membrane storage part 20 to react with hemoglobin collected therein (more precisely, hemoglobin with metal ions bound thereto) May be transferred to the buffer solution storage part 30 to react with the buffer solution.

The conversion material storage unit 40 includes a conversion material that converts the reacted hemoglobin into methylated hemoglobin (Met Hb) by reacting with the hemoglobin from which the metal ion is separated. That is, the conversion substance is a substance that converts various types of hemoglobin into methylated hemoglobin, and can convert all forms of hemoglobin hemoglobin, hemoglobin, and carbon monoxide into hemoglobin, such as cyanide, sodium azide, Sodium Dodecyl Sulfate, and the like. Furthermore, for their stability, the conversion material can further comprise a surfactant, and when present in a solidified solid phase, the stability of the product can be further enhanced, and when converted to a liquid phase upon reaction with hemoglobin, desirable. In the present invention, all the hemoglobin of various types can be measured through such a conversion material, and more accurate hemoglobin amount detection is possible. The conversion material is transferred from the conversion material storage unit 40 to the membrane storage unit 20 (or the buffer solution storage unit 30), and the hemoglobin present therein (more precisely, the hemoglobin from which the metal ions are separated) It is also possible that the hemoglobin collected in the membrane storage part 20 (or the buffer solution storage part 30) is transferred to the conversion substance storage part 40 to react with the conversion substance.

The absorbance measurement means (not shown) measures the absorbance of the methylated hemoglobin present in the conversion substance storage section 40 (or the membrane storage section 20 or the buffer solution storage section 30) . That is, it is possible to measure the absorbance using a light source of a multi-wavelength band in a place where the hemoglobin is methylated by reacting with the conversion substance. The methylated hemoglobin may be present in the conversion substance storage unit 40 (or the membrane storage unit 20 or the buffer solution storage unit 30), and the absorbance measurement unit according to the present invention may be provided in each storage unit 20 30, and 40, and may be provided over a region including all of the storage units 20, 30, and 40, for example.

For example, the hemoglobin (Hb) of hemolyzed blood cells has a porphyrin structure as described above, which shows a characteristic absorbance in the region of 413, 540 and 575 nm. In order to detect hemoglobin more accurately, the absorbance at the reference wavelength band 800 nm is used together with the absorbance at 413, 540 and 575 nm, which are the porphyrin characteristic wavelength bands, together with the detection of the amount or concentration of total hemoglobin using these absorbance values. Thus, more accurate hemoglobin concentration can be calculated based on the signal of total hemoglobin (S_total Hb) through the linear relationship. In order to improve the accuracy of total hemoglobin detection, when the wavelength of the relative radiant intensity (RRI) of the LED is 0.5 or more as a signal source, data of about 50 nm or longer is received by the CCD and subjected to Multivariate linear regression ) Can be applied. The present invention can detect the total amount of hemoglobin necessary for the calculation of the glycated hemoglobin concentration through the absorbance using the absorbance measuring means.

The oxidase storage unit 50 includes an oxidase which is mixed with hemoglobin from which the metal ions are separated. That is, the oxidase is a substance for measuring glucose-bound hemoglobin in the hemoglobin, for example, glucose oxidase (GOx). The oxidizing enzyme is present only in a mixed state with the hemoglobin containing the glycated hemoglobin and reacts with a signal generating substance described later on the electrode unit 60 to generate an electric signal, . ≪ / RTI > The oxidase storage part 50 includes the oxidizing enzyme in advance and the hemoglobin in which the metal ions are separated by reacting with the buffer solution is injected into the oxidizing enzyme storage part 50 so that the mixture of the oxidizing enzyme and the hemoglobin isolated from the metal ion Lt; / RTI > The present invention is characterized in that the oxidase is contained in the separate oxidase storage part 50 and the hemoglobin which reacts with the oxidase is hemoglobin in which metal ions are separated. The hemoglobin in which the metal ions are separated minimizes the influence of other interference substances for the measurement of glycated hemoglobin. Since the hemoglobin is methylated by the conversion substance, the reaction for measurement of glycated hemoglobin is started before or after the measurement of the hemoglobin amount So that it is preferable.

The electrode unit 60 is immobilized with a sugar solution capable of binding with the sugar of the hemoglobin and the oxidase by reacting with the mixture of the hemoglobin and the oxidase in which the metal ions are separated. That is, the electrode unit 60 is for measuring the amount of hemoglobin of glycoprotein by an electrochemical method, and sugar binding means (or glycated hemoglobin coupling means) is immobilized in advance therein. Among the hemoglobin, the sugar-binding means may be Boronic Acid which recognizes or binds sugar of hemoglobin and oxidase, or may be an antibody of HbA1c, and may be a self-assembly of Cys-FPBA or Dend-BA A self assembly monolayer (SAM) is preferable. Since both HbA1c and GOx are glycoproteins, it is possible to immobilize the surface with modified cis-diol interactions on the surface modified with voronic acid without any pretreatment. As described above, when the sugar-binding means is immobilized on the electrode unit 60, and the mixed solution in which the hemoglobin and the oxidase in which the metal ions are separated is introduced into the electrode unit 60, the sugar- It is coupled with an enzyme to prepare to generate an electrical signal.

The signal-generating material storage unit 70 includes a signal generating material for generating an electrical signal by reacting with hemoglobin and an oxidizing enzyme attached to the electrode unit. That is, the signal generating material generates an electric signal according to the amount of glycated hemoglobin in the electrode part, for example, ferrocene and glucose. The glucose reacts with the above-mentioned oxidase (GOx), and the ferrocyan generates electric signals in this process, and it can be ferrocyanide and / or ferri cyanide Do. The present invention is to measure the amount of the glycated hemoglobin using a competitive binding assay of glycated hemoglobin and oxidase. The concentration of the glycated hemoglobin is high, and as the gel is immobilized on the surface, The amount of the oxidizing enzyme immobilized is also small and the signal is reduced. The signal generating material may further include a conductivity enhancing material including HAuCl ₄ and a nicotinamide adenine dinucleotide (NAD), and may further include SDS (sodium dodecyl sulfate) It is also possible to include it.

The electrical signal measuring means (not shown) measures an electrical signal generated by the electrode unit 60 and measures the amount of HbA1c. That is, the electrical signal measuring means measures an amount of HbA 1c by an electrochemical method by receiving an electrical signal that varies depending on the amount of HbA 1c in the electrode unit 60. For example, a glycated hemoglobin detection signal can acquire a redox curve by a continuous cyclic voltametry technique. The percentage of glycated hemoglobin can be obtained by calculating the percentile of the total hemoglobin concentration using the current value obtained at a specific voltage within the curve. In addition, it is possible to quantitatively obtain the amount of glycated hemoglobin by detecting an electrochemical signal of glycated hemoglobin using an electrochemical detection method such as a chronoamperometry and a differential partial voltametry method .

The present invention as described above can more accurately calculate the glycated hemoglobin concentration by separately measuring the porphyrin absorbance signal inherent to hemoglobin (including glycated hemoglobin) and the electrochemical signal by the oxidase separately. In addition, the present invention can improve the accuracy of detection by separating only hemoglobin (especially hemoglobin in which metal ions are removed) from the blood, and enables miniaturization and low-cost system according to electrochemical signal detection.

Further, the present invention is characterized in that a reaction solution or a reaction substance necessary for the detection of glycated hemoglobin is independently implemented. Thus, it is possible to prevent the solution from remaining in the channel without mixing with the reaction solution, It is possible to more accurately measure the content of hemoglobin and the glycated hemoglobin by preventing the overflow. In addition, the present invention can minimize the influence of errors generated by driving each reaction solution of the cartridge in a common channel and other interference substances in the blood, by individually controlling each fluid such as a reaction solution or a sample There is an effect.

FIG. 4 is a perspective view showing an example of the configuration of the fluid transferring means according to the present invention, and FIG. 5 is a schematic view for explaining an example of a state in which the fluid transferring means according to the present invention is moved to each storage portion of the cartridge through the position sensing means .

As described above, according to the present invention, it is possible to move the reaction liquid and the like contained in each of the storage portions through the separate fluid transfer means 100. When the fluid transfer means 100 is a general syringe, the user can manually move the reaction fluid using the fluid transfer means 100. When the fluid transfer means 100 is mounted on another automatic transfer means, As shown in FIG.

As shown in FIG. 4, the fluid transfer means 100 may include a fluid loading unit 120 for loading fluid and a syringe 110 for loading the fluid. The fluid loading part 120 is made up of a thin tube and is capable of introducing fluid such as blood, sample, reaction liquid, etc. into the inside thereof by the micro pressure, and the fluid introduced by the pressure of the syringe 110 . A driving pipe may be further included between the fluid loading unit 120 and the syringe unit 110 to transfer the fluid. In addition, the fluid loading unit 120 may previously contain blood for hemolysis of the blood sample before the initial blood sample is introduced.

The fluid transfer means 100 may include at least one selected from the group consisting of the metal ion storage portion, the membrane storage portion, the buffer solution storage portion, the conversion material storage portion, the oxidizing enzyme storage portion, the electrode portion, and the signal generating material storage portion Or one or more fluids. The fluid may be transferred from one storage unit to another storage unit selected from the group consisting of the metal ion storage unit, the membrane storage unit, the buffer solution storage unit, the conversion substance storage unit, the oxidizing enzyme storage unit, the electrode unit, It is also possible to do. The path to be transferred to each storage unit will be described in detail in the detection method to be described later.

In addition, the fluid transfer means 100 senses a position corresponding to each of the metal ion storage portion, the membrane storage portion, the buffer solution storage portion, the conversion substance storage portion, the oxidizing enzyme storage portion, the electrode portion, And a position sensing unit 130 for sensing the position of the object. The position sensing unit 130 includes an LED, a photodiode, and / or an ultrasonic sensor for sensing the x-axis, the y-axis, and / or the z-axis, respectively. Thus, as shown in Fig. 5, it is possible to recognize a calibration located at a specific position on the cartridge or around each storage unit, and automatically move along a predetermined path while exchanging signals with it.

To this end, the present invention may further include a control unit (not shown) for automatically moving the fluid transfer means 100 according to a signal received from the position sensing means 130.

6 is a flowchart illustrating a process of a method for detecting glycated hemoglobin according to another embodiment of the present invention.

The method of detecting glycated hemoglobin according to the present invention can be carried out using the cartridge-type glycated hemoglobin detecting apparatus as described above.

The present invention is to detect the concentration ratio of glycated hemoglobin among hemoglobin in blood cells, and it is preferable to first hemolyze red blood cells which occupy about 50% in the blood. Methods for hemolysis include a method using an osmotic imbalance of hypo-osmolarity or hyper-osmolarity, a method using pH control of Hypo-tonic or Hyper-tonic, or a method using high- (High Shear Stress) or mechanical stress caused by a vibration field, or may be hemolyzed using thermal stress at a temperature of 48 ° C or higher. For example, when whole blood (1 to 5 ㎕) is injected into the fluid transfer means of the present invention, hemolysis is carried out in a tube containing the blood for the fluid transfer means, and a sample in which blood cells are separated from the plasma can be prepared (S1).

Next, the present invention includes a step (S10) of injecting a sample into the metal ion storage portion 10 containing metal ions capable of binding hemoglobin (Hb). The metal ion storage unit 10 is as described above. That is, the sample is reacted with the metal ion by injecting the sample into the metal ion storage portion 10 using the fluid transfer means. By the reaction, the metal ion storage part 10 may include a sample containing hemoglobin in which metal ions are bound.

In this case, the fluid transferring means may include a sample remaining after being injected into the metal ion storage portion 10, and the method of the present invention may further include a step of discarding the surplus fluid remaining in the waste solution storage portion 80 And the apparatus of the present invention may further include a first waste solution storage part 81 in which the excess liquid can be contained. In this way, the remaining surplus liquid can be contained together in the cartridge of the present invention, and a batch waste solution treatment is possible.

Next, the step of injecting a sample reacted with the metal ion into a membrane storage section 20 including a membrane separating the hemoglobin bound to the metal ion and the material not bound to the metal ion (S20 ). That is, the sample stored in the metal ion storage unit 10 is transferred to the membrane storage unit 20 through the fluid transfer unit. Then, the sample is separated into the hemoglobin bound to the metal ion and the material not bound to the metal ion by the membrane 21 included in the membrane storage part 20.

Herein, it is preferable that the substance not bound to the metal ion is discharged to the outside of the membrane storage part 20. In the method of the present invention, the substance not bound to the metal ion is separated from the waste solution storage part 80, The apparatus further includes a second waste solution storage part 82 in communication with the membrane storage part 20, in which a substance not bound to the metal ion is moved and stored, ). ≪ / RTI > When the sample is injected into the membrane storage part 20, a substance not bound to the metal ion passes through the membrane and is naturally transferred to the second waste solution storage part 82. Through this, it is possible to improve the accuracy of detection by removing interfering substances which are unnecessary or disturbing in hemoglobin measurement, and the interfering substances can be contained together in the cartridge of the present invention, and batch waste solution treatment is possible.

In addition, the fluid transferring means may include a sample (a sample containing hemoglobin bound to metal ions) injected into the membrane storage portion 20, and the method of the present invention may use the washing liquid storage portion 90 The apparatus of the present invention may further include a first washer liquid storage unit 91 for washing the sample. That is, according to the present invention, the inside of the fluid transfer means can be cleaned through a process of moving the fluid transfer means injected with the sample to the first washer liquid storage portion 91 and suctioning the fluid once more. Thus, more accurate results can be obtained.

Next, the method of the present invention is characterized in that the buffer solution is introduced from a buffer solution storage part 30 containing a buffer solution for separating hemoglobin bound to metal ions into metal ions and hemoglobin, and injected into the membrane storage part 20 (S30). That is, the buffer solution stored in the buffer solution storage unit 30 is transferred to the membrane storage unit 20 through the fluid transfer unit. Then, the hemoglobin bound to the metal ion by the buffer solution is separated into metal ions and hemoglobin.

Thereafter, the present invention goes through a step (S40) of injecting the hemoglobin into which the metal ions have been separated by the buffer solution into a conversion substance storage section 40 containing a conversion substance which converts the hemoglobin into methylated hemoglobin (Met Hb). That is, the hemoglobin (hemoglobin in which the metal ions are separated) stored in the membrane storage unit 20 is transferred to the converted material storage unit 40 through the fluid transfer unit. Then, the hemoglobin is converted into methylated hemoglobin by the conversion substance.

Next, the present invention includes an absorbance measurement step (S50) of measuring the absorbance of the hemoglobin of the methylated hemoglobin present in the converted substance storage section (40) and measuring the hemoglobin amount. That is, the total hemoglobin amount can be measured in the converted substance storage section 40 where the methylated hemoglobin exists using the absorbance measurement means.

Next, in the present invention, the hemoglobin from which the metal ions are separated by the buffer solution is injected into the oxidase storage part 50 including the oxidase (S60). This step may be performed before, simultaneously with, or after the step (S40) of injecting the hemoglobin into which the metal ions have been separated into the conversion substance storage section 40. Since the same hemoglobin is transferred, it is more preferable that it is continuous. Therefore, the oxidase storage unit 50 may contain a mixture of hemoglobin and oxidase in which the metal ions are separated.

In this case, the fluid transfer means transferring the hemoglobin from which the metal ions have been separated may contain the sample remaining after being injected into the conversion substance storage section 40 and the oxidizing enzyme storage section 50, (S45) of washing the sample using the storage unit 90, and the apparatus of the present invention may further include a second washer liquid storage unit 92 for washing the sample . That is, according to the present invention, the inside of the fluid transfer means can be cleaned through a process in which the fluid transfer means injected with the sample is moved to the second washer liquid storage portion 92 and suctioned at least once, Thus, more accurate results can be obtained.

(S70) injecting a mixture of hemoglobin and oxidase, in which the metal ions are separated, into an electrode unit 60 to which sugar binding means capable of binding with the hemoglobin and the oxidase are immobilized, ≪ / RTI > That is, the mixed solution (mixed liquid of hemoglobin and oxidase enzyme from which metal ions have been separated) is transferred to the electrode unit 60 through fluid transfer means. Then, the hemoglobin and the oxidizing enzyme may be bound to the sugar-binding means.

In the method of the present invention, the substance not coupled to the sugar-binding means is separated from the waste solution storage part 80, The apparatus further includes a third waste solution storage part 83 which is connected to the electrode part 60 and in which a substance not associated with the sugar binding means is moved and stored, ). ≪ / RTI > When the mixed solution is injected into the electrode unit 60, a substance which can not be combined with the sugar-binding unit is naturally moved to the third waste solution storage unit 83 and discharged. Through this, it is possible to improve the accuracy of detection by removing interfering substances which are unnecessary or disturbing in hemoglobin measurement, and the interfering substances can be contained together in the cartridge of the present invention, and batch waste solution treatment is possible.

In the present invention, the signal generating material is introduced from the signal generating material storing unit 70 containing a signal generating material which reacts with hemoglobin and oxidizing enzyme to generate an electric signal, and is injected into the electrode unit 60 Step S80 is performed. That is, the signal generating material stored in the signal generating material storage unit 70 is transferred to the electrode unit 60 through the fluid transferring unit. Then, the signal generating material may generate a signal depending on the amount of glycated hemoglobin in the electrode unit 60. In this case, it is preferable that the signal generating material is injected in an amount larger than the hemoglobin and the amount of the oxidizing enzyme, preferably 2 to 4 times, so that sufficient electrochemical reaction can be performed, Because substances (GOx, Hb) and interfering substances (SAM constituent residues) can be used as wash liquids and / or electron transfer mediators.

Furthermore, the present invention is characterized by measuring an electrical signal generated by the electrode unit 60 and measuring an amount of HbA1c (S90). That is, it is possible to measure the amount of glycated hemoglobin from the electrical signal generated by the electrode unit 60 using the electrical signal measuring means, and to calculate the glycated hemoglobin concentration accordingly.

FIG. 7 is a graph showing the results of measurement of hemoglobin content in an optical system according to an embodiment of the present invention, and FIG. 8 is a graph showing the results of measurement of a glycated hemoglobin content in an electrochemical manner according to an embodiment of the present invention to be.

That is, FIG. 7 shows the total hemoglobin value measured through the optical signal by the optical method, and it can be seen that the optical signal differs according to the hemoglobin concentration.

8 is a graph showing a current value according to the concentration of glycated hemoglobin when sweeping at a scan rate of 5 mV / s by applying a cyclic voltammetry method in an electrochemical manner. The percentage of glycated hemoglobin can be obtained by calculating the glycated hemoglobin value using the current value obtained at a specific voltage and calculating the percentile value of the total hemoglobin concentration using the total hemoglobin value measured by the optical signal there was.

While the present invention has been particularly shown and described with reference to preferred embodiments thereof, it is to be understood that the invention is not limited to the disclosed embodiments, but, on the contrary, It will be apparent to those skilled in the art that changes may be made.

10: metal ion storage part
11: Metal ion
20: Membrane storage part
21: Membrane
30: Buffer liquid storage unit
40: conversion material storage unit
50: oxidase storage
60:
70: Signal generating substance storage unit
80: waste solution reservoir
81: First waste solution storage part
82: Second waste solution storage part
83: Third waste solution storage part
90: Washing liquid storage part
91: First washer fluid storage part
92: second washer liquid storage part
100: fluid transport means
110: Cirin Branch
120: fluid loading section
130: Position sensing means

Claims (15)

delete delete delete delete delete delete delete delete delete delete delete The detection of glycated hemoglobin using a cartridge-type glycated hemoglobin detecting device including a metal ion storage part, a membrane storage part, a buffer solution storage part, a conversion substance storage part, an oxidase storage part, a signal generating substance storage part, an electrode part and a fluid transferring part In the method,
(S1) preparing whole blood and blood plasma separated sample by injecting whole blood into the fluid transfer means including blood for blood;
(S10) injecting the sample into a metal ion storage portion containing metal ions capable of binding hemoglobin (Hb);
(S20) injecting a sample reacted with the metal ion into a membrane storage part including a membrane separating the hemoglobin bound to the metal ion and the material not bound to the metal ion;
(S30) a step of injecting the buffer solution into the membrane storage part from a buffer solution storage part containing a buffer solution for separating hemoglobin bound to metal ions into metal ions and hemoglobin;
(S40) injecting a hemoglobin into which a metal ion has been separated by the buffer solution into a conversion substance storage section containing a conversion substance which converts the hemoglobin into methylated hemoglobin (Met Hb);
An absorbance measurement step (S50) of measuring the absorbance of the hemoglobin of methylated hemoglobin present in the conversion substance storage section and measuring the hemoglobin amount;
Injecting hemoglobin into which the metal ions have been separated by the buffer solution into an oxidase storage unit containing the oxidase (S60);
(S70) injecting a mixed solution in which the hemoglobin and the oxidase are separated from the metal ion into an electrode unit to which sugar binding means capable of binding with the hemoglobin and the oxidizing enzyme is immobilized;
A step S80 of injecting the signal generating material into the electrode unit from a signal generating material storing unit including a signal generating material which reacts with hemoglobin and oxidizing enzyme to generate an electric signal;
And an electrical signal measuring step (S90) of measuring an electrical signal generated from the electrode unit and measuring an amount of HbA1c,
The storage units are divided into separate independent spaces so as not to communicate with each other. The storage units are each provided with a hole through which the fluid can be introduced into the storage units,
Wherein the fluid transferring unit includes a fluid loading unit for loading fluid and a syringe unit for loading the fluid, and the fluid transferring unit includes a syringe unit for loading the fluid, Wherein the movement of the sample, the buffer solution, the hemoglobin, the mixed solution, or the signal generating material is performed by the fluid transfer means,
The step (S10) may further include the step of discarding the remaining sample in the first waste solution storage section when the fluid transfer means includes a sample remaining after being injected into the metal ion storage section,
The method of any one of the preceding claims, further comprising the step of discharging a substance not bound to the metal ion through the membrane to a second waste solution reservoir communicated with the membrane reservoir,
The method of claim 1, further comprising the step of discharging a substance not bound to the sugar binding means to a third waste solution storage portion communicating with the electrode portion.
13. The method of claim 12,
Wherein the conversion material contained in the conversion material storage unit is solidified.
13. The method of claim 12,
The fluid transfer means includes position sensing means for sensing a position corresponding to each of the metal ion storage portion, the membrane storage portion, the buffer solution storage portion, the conversion material storage portion, the oxidizing enzyme storage portion, the electrode portion, and the signal generating material storage portion Further comprising the step of detecting the glycated hemoglobin.
15. The method of claim 14,
Wherein the cartridge-type glycated hemoglobin detecting apparatus further comprises a controller for automatically moving the fluid transferring means in accordance with the position sensing means, thereby automatically moving the fluid transferring means.

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