KR101121005B1 - Bio chip including phase change layer, method of detecting bio chip using phase change and diagnostic kit with the bio chip - Google Patents

Abstract

The present invention relates to a biochip comprising a phase change layer, a biochip detection method using a phase change, and a diagnostic kit including the biochip, wherein the biochip includes a substrate, an electrode, a phase change layer, a protective film, an organic material, and a nanoparticle. In the chip, the phase change layer is a chalcogenide composed of one or more elements selected from the group consisting of Te, Se, Ge, Sb, Bi, Pb, Sn, As, In _, Ag, S, Si, P, and O. It is characterized by the use of a substance.

According to the present invention, since the resistance between the two electrodes connected to both ends of the material in which the phase change occurs can be detected, it is possible to provide a biochip that can easily detect target probes such as genes, By simplifying the structure of the biochip, it is possible to provide a miniaturized device and a possible detection method and diagnostic kit at low cost.

Phase change layer, bio chip, electrode, probe, organic material, electrical properties, diagnostic kit

Description

BIO CHIP INCLUDING PHASE CHANGE LAYER, METHOD OF DETECTING BIO CHIP USING PHASE CHANGE AND DIAGNOSTIC KIT WITH THE BIO CHIP}

The present invention relates to a biochip including a phase change layer, a biochip detection method using a phase change, and a diagnostic kit including a biochip, and more specifically, to easily diagnose a gene using a phase change phenomenon of a phase change layer. The present invention relates to a biochip including a phase change layer that can be easily and easily used, a method for detecting a biochip such as a gene using the biochip, and a diagnostic kit including the biochip.

A biochip is a chip in which biomolecules probes such as DNA and protein to be attached on a substrate are densely attached, and the gene expression patterns, gene defects, protein distribution, and reaction patterns in a sample can be analyzed. Biochips can be divided into microarray chips attached to a solid substrate and lab-on-a-chip attached to the microchannels according to the attachment form of the probe.

In such a biochip, a system capable of detecting whether a target molecule is bound to a probe immobilized on a substrate is required to determine whether a target molecule capable of binding a probe exists in a sample.

Currently, DNA chips for genetic analysis mostly label fluorescent dyes on sample DNA, react with probes on the chip, and detect fluorescent materials remaining on the chip surface using a confocal microscope or CCD camera. Use the method.

However, since such an optical detection method is difficult to miniaturize and the digitized output cannot be seen, much research is being conducted on the development of a new detection method capable of producing an electrical signal.

Many research institutions, including the Clinical Micro Sensor, are investigating how to detect DNA hybridization electrochemically using metal compounds that are easy to oxidize / reduce. When DNA hybridizes, other compounds, including metals that are easily oxidized / reduced, form a complex together and are detected electrochemically. However, the electrochemical method also has the disadvantage of requiring separate labeling.

In addition, research is being actively conducted on the analysis without using fluorescent dyes or any other markers. A method for measuring mass difference before and after binding using quartz crystal microbalance, and matrix assisted laser adsorption ionization Methods of analysis using mass spectrometry have been developed.

In addition, up to one base difference could be analyzed using a microfabricated cantilever, a mechanical sensor type that measures the intermolecular binding force before and after binding of a DNA probe and a target.

However, in this case, the deflection of the cantilever beam must be measured very precisely. For this purpose, additional equipment such as a laser is required.

Applicant to the above problems by using the phase change layer, by measuring the resistance between the electrodes connected to both ends of the phase change layer to detect the crystallization of the phase change layer, it is possible to simply and accurately detect the gene, etc. Although a biochip detection method has been filed and registered, it discloses a technique for a conceptual and overall level of detection method in which a specific technology regarding a phase change layer or an electrode, which is a component of a biochip, is not disclosed.

The present invention has been made to solve the above problems, by detecting the resistance between the two electrodes connected to both ends of the phase change layer including a bio-spot where the phase change occurs, detection of the binding of the biochip It is an object of the present invention to provide a biochip that can be detected using a phase change.

Another object of the present invention relates to a method for detecting using the biochip.

Still another object of the present invention is to provide a diagnostic kit including the biochip.

The present invention is to achieve the above object, in the biochip comprising a substrate, an electrode, a phase change layer, a protective film, an organic material and nanoparticles, the phase change layer is Te, Se, Ge, Sb, Bi, Pb It provides a bio-chip comprising a phase change layer, characterized in that using a chalcogenide material consisting of at least one element selected from the group consisting of, Sn, As, In _, Ag, S, Si, P and O.

In addition, the phase change layer is Ge _a Sb _b Te _c (a, b, c is the atomic mole fraction, a + b + c = 1, 0 <a, b, c <1) or In _d Ag _e Sb _f Te _g (d, e, f, g is characterized by using an atomic mole fraction, d + e + f + g = 1, 0 <d, e, f, g <1), respectively.

In addition, the electrode is titanium (Ti), titanium nitride (TiN), tungsten (Tungsten, W), gold (Au), aluminum (Aluminum, Al), tantalum (Tantalum, Ta), tantalum nitride (TaN) ), Molybdenum (Molybdenum, Mo) and copper (Copper, Cu) is characterized in that it comprises a metal selected from.

The protective film may be an oxide film or a nitride film.

In addition, the organic material is characterized in that the DNA, PNA, protein or pigment.

In addition, the organic material is characterized in that it comprises a fixed probe fixed to the phase change layer; and a target probe capable of complementarily coupling to the fixed probe.

In addition, the nanoparticles are characterized in that coupled to the target probe.

In addition, the protective film is characterized in that the surface treatment.

In addition, the present invention provides a biochip detection method using the biochip, comprising: preparing a biochip in which a fixed probe, which is an organic material, is fixed to a biospot of a phase change layer connected between the electrodes; Bonding the fixed probe to the biochip by spraying a target probe having a nanoparticle bound to the biochip; Irradiating laser light onto the biospot; Detecting a phase change by measuring resistance between the electrodes, wherein the phase change layer comprises Te, Se, Ge, Sb, Bi, Pb, Sn, As, S, Si, P, and O It provides a biochip detection method using a phase change, characterized in that using a chalcogenide material consisting of one or more elements selected from.

The present invention also provides a diagnostic kit comprising the biochip.

According to the biochip including the phase change layer of the present invention, since the resistance between both electrodes connected to both ends of the material in which the phase change occurs can be detected, it is possible to facilitate detection of target probes such as genes. The biochip can be provided, and the structure of the biochip can be simplified to provide a small sized device and a detection method and a diagnostic kit at low cost.

The present invention is a biochip comprising a substrate, an electrode, a phase change layer, a protective film, an organic material and nanoparticles, the phase change layer is Te, Se, Ge, Sb, Bi, Pb, Sn, As, In _, Ag It relates to a biochip comprising a phase change layer, characterized in that using a chalcogenide material consisting of at least one element selected from the group consisting of S, Si, P and O.

Hereinafter, the present invention will be described in detail with reference to the accompanying drawings.

Biochip of the present invention composed of the above components, from the bottom, the substrate, the electrode formed on the substrate, the phase change layer formed on the electrode, the protective film formed thereon, the organic material formed thereon and nanoparticles attached to the organic material consist of.

In addition, the order of the components may be selectively changed, and sequentially from below, the substrate, the phase change layer formed on the substrate, the electrode formed on the phase change layer, the protective film formed thereon, the organic material formed thereon and nano-attached to the organic material It may also consist of particles.

The substrate serves to facilitate electrode fabrication, surface treatment, or organic material adhesion, and may use chromium glass, silicon wafer, polymer, and the like, and has a non-conductive property. If it is, the conventionally used one can be used without limitation.

The phase change layer is a chalcogenide material consisting of at least one element selected from the group consisting of Te, Se, Ge, Sb, Bi, Pb, Sn, As, In _, Ag, S, Si, P and O, Ge _a Sb _b Te _c (a, b, c are atomic mole fractions, a + b + c = 1, 0 <a, b, c <1) or In _d Ag _e Sb _f Te _g (d, e, f, g Uses atomic mole fraction, d + e + f + g = 1, 0 <d, e, f, g <1), respectively, and it is particularly preferable to use Ge ₂ Sb ₂ Te ₅ . Phase change layer is a thermal change such as laser irradiation, when there is a change over a certain temperature, the crystallization is made of a material that causes a phase change, the biochip of the present invention uses the characteristics of such a phase change layer To detect genes.

The electrode is connected to both sides of the phase change layer and serves to detect the phase change of the phase change layer by measuring the resistance between both electrodes, and according to the degree of phase change in the phase change layer, As the fraction increases, the resistance is lower than the reference resistance. The higher the fraction is, the higher the resistance becomes than the reference resistance.

The metal used as the electrode is titanium (Ti), titanium nitride (TiN), tungsten (W), gold (Au), aluminum (Aluminum, Al), tantalum (Tantalum, Ta), tantalum nitride (TaN), molybdenum (Molybdenum, Mo), copper (Copper, Cu) may be exemplified, and can be used without limitation as long as it has conductivity characteristics.

The protective film serves to prevent surface short and vaporization of the phase change layer between devices and to enable surface treatment for raising DNA, and is transparent and can be used without limitation as long as it has the characteristics of an insulator. An oxide film and a nitride film can be used. Particularly, it is preferable to use silicon oxide (SiO ₂ ) in the oxide film, since the surface treatment for DNA adsorption used in conventional slide glass is possible.

The protective film is subjected to a surface treatment including the following steps.

(a) attaching an OH group to the surface of the protective film;

(b) treating (3-aminopropyl) trimethoxysilane (APTMS) on the OH group; And

(c) attaching the aldehyde

The reason for the surface treatment in this way is to attach the organic material to be subsequently processed at a high density.

The organic material includes a fixed probe fixed to the phase change layer and a target probe that can be complementarily bound to the fixed probe. As the type thereof, DNA, PNA, protein, or dye may be used depending on a target to be detected. .

The nanoparticles are attached to the target probe, so that the presence of the nanoparticles blocks the laser light, which causes the phase change of the phase change layer, and thus, the presence of the target probe can be detected by preventing the phase change from occurring. To ensure that

As the mechanism, depending on the degree of phase change in the phase change layer, the higher the crystalline fraction, the lower the resistance than the predetermined reference resistance, and the higher the amorphous fraction, the higher the reference resistance. It is to show resistance and the reference resistance can be determined by experiment.

As the nanoparticles, metals such as gold, silver, and copper may be used.

The present invention also relates to the following biochip detection method.

(a) preparing a biochip in which a fixed probe, which is an organic material, is fixed to a biospot of a phase change layer connected between electrodes;

(b) sprinkling the target probe with the nanoparticles bound to the ends of the biochip, thereby binding to the fixed probe;

(c) irradiating laser light to the biospot;

(d) detecting the phase change by measuring the resistance between the electrodes

The phase change layer is a chalcogenide material consisting of at least one element selected from the group consisting of Te, Se, Ge, Sb, Bi, Pb, Sn, As, In _, Ag, S, Si, P and O, Ge _a Sb _b Te _c (a, b, c are atomic mole fractions, a + b + c = 1, 0 <a, b, c <1) or In _d Ag _e Sb _f Te _g (d, e, f, g Uses the atomic mole fraction, d + e + f + g = 1, 0 <d, e, f, g <1), respectively. Since other components of the biochip are the same as described above, they are omitted.

The present invention also relates to a diagnostic kit which can easily diagnose a disease by easily detecting a gene or the like using the biochip. The diagnostic kit includes the biochip and data collection and diagnostic equipment connected thereto, which may be in the form of a diagnostic kit used in a large hospital. Also, the biochip may be connected to a portable device for personal use at home. It can also be used in the form of a diagnostic kit.

1 is a diagram schematically illustrating a system configuration capable of detecting electrical characteristics of a biochip including a phase change layer. The portable biochip scanner connects and communicates with a PC via USB, and uses a laser to crystallize the phase change layer and inserts the biochip scanner into a biochip scanner to provide a voltage (-1V to + +). 1V, user-definable) to detect the resistance of the biochip.

On the other hand, raw data of a resistance signal detected by a biochip in a PC may be converted into an image. Accordingly, the biochip information may be detected through the biochip image conversion software and analysis software for analyzing and reporting the biochip image.

Hereinafter, the present invention will be described in detail with reference to Examples. The following examples are provided to illustrate the present invention, and the scope of the present invention includes the invention described in the claims below, its modifications and substitutions, and is not limited to the scope of the following examples.

[Example]

Phase change layer  Biochip Fabrication Including

(One) Ti Of TiN  Unit electrode formation process

Referring to FIGS. 2 and 3, a unit electrode forming process will be described. A MASK (not shown) is manufactured by using the LAY-OUT shown in FIGS. 2 and 3, and then, the MASK is used. Proceed with the recorded procedure of Table 1 to produce the unit electrode (FABRICATION).

Referring to FIG. 4, the process will be described in detail. A first insulating film is formed on a Si wafer. Preferably, the first insulating film is thermal oxide. Ti / TiN is then deposited in-situ. Next, a lithography (LITHOGRAPHY) process is performed using the MASK. Here, an ashing process is performed using N ₂ / Ar gas in an ICP method to improve the degree of removal of the photoresist. As a result, a first preliminary biochip having a Ti / TiN unit electrode is manufactured.

fair Contents Remarks Oxide Thermal oxide 1000 nm Metal Deposition Ti / TiN Ti 5nm / TiN 25nm, sputter deposition lithography


Photo process PR Coating, align, development Etch Process Use RIE method Strip process Use PR REMOVER Ashing N ₂ / Ar gas, ICP ASHING

(2) GST ( Ge ₂ Sb ₂ Te ₅ ) / SiO ₂ layer  Forming process

A GST / SiO ₂ layer is formed on the first preliminary biochip. Specifically, referring to FIG. 5, the GST and SiO ₂ layer deposition equipment is designed to deposit GST and SiO ₂ films by using a magnetron sputtering method. The GST to be deposited here is Ge ₂ Sb ₂ Te ₅ , a chalcogenide compound, and the process conditions are Base Pressure <5.0X10 ^-6 Torr, Power = DC 10 W, and the like. In order to deposit 30 GST, Deposition time = 58 sec.

And 20 nm of SiO ₂ is deposited on the GST in-situ. At this time, the process conditions for SiO ₂ deposition are Base Pressure <5.0X10 ^-6 Torr, Power = RF 150 W. In order to deposit SiO ₂ 20 nm, Deposition time = 1min 58sec. As a result, a second preliminary biochip having a GST / SiO ₂ layer formed on the Ti / TiN unit electrode is manufactured.

(3) SiO ₂  Surface treatment process

Specifically, referring to FIG. 6, a surface treatment is performed on the OXIDE surface in order to subsequently attach organic materials and nanoparticles at high density onto the second preliminary biochip. The surface treatment is performed in three stages: OH treatment, APTMS ((3-aminopropyl) trimethoxysilane) treatment, and aldehyde (Glutaradehyde) treatment. Each process is described as follows.

(3-1) OH treatment

OH treatment on the surface of the second preliminary biochip using O ₂ Plasma. Maintain a base pressure of 50 Pa in the sputtering chamber during surface treatment. After setting the flow rate of oxygen to 300 ml / min, the RF output is set to about 500 Watts and the surface is treated for about 2 minutes.

(3-2) (3-aminopropyl) trimethoxysilane (APTMS) Treatment

After the OH treatment described above, it is then sufficiently washed with DI water and ethanol. The substrate is immediately immersed in APTMS solution (1%) without drying and placed in a sealed container for 1 hour at room temperature. After storage, the substrate is thoroughly washed with ethanol and dried with gaseous nitrogen. The substrate is then placed on a hotplate at 150 ° C. for 1 hour.

(3-3) Aldehyde (-CHO) surface treatment

After the above-mentioned APTMS treatment, and then immersed in glutaraldehyde solution (2%) and put in a sealed container and stored at room temperature for 2-4 hours. After this step, wash with DI water and dry with nitrogen gas. As a result, a third preliminary biochip subjected to surface treatment is manufactured.

(4) Fixed probe DNA  And labeled with nanoparticles target DNA  Attachment technology process

Referring to FIG. 7, the organic material and the nanoparticles are attached onto the third preliminary biochip. The attachment process consists of a fixed probe DNA attachment process and a target DNA attachment process represented by nanoparticles. Each adhesion process is demonstrated as follows.

(4-1) Fixed Probe DNA Attachment Step

First, the third preliminary biochip is washed with 0.2% SDS for 10 minutes. Subsequently, the third preliminary biochip is washed twice with tertiary distilled water for 5 minutes. Next, the reaction was stirred for 10 minutes in a solution of 0.7 g sodium borohydride and 500 ml reduction solution. The mixed solution should be prepared just before use. Then wash for 5 minutes with tertiary distilled water. Then wash for 5 minutes with 0.2% SDS. Then, washed three times with 1 minute distilled water. Then dry using N ₂ gas or centrifuge. The dried third preliminary biochip is then placed in a slide box containing a moisture absorbent and stored in a desiccator. As a result, the fixed probe DNA is attached onto the third preliminary biochip.

(4-2) labeled with nanoparticles target DNA  Attachment process

The process for attaching the target DNA labeled with nanoparticles to the fixed probe DNA described above is described below.

First, 5 μl of Mycobacteria Genotyping PCR Product, streptavidin conjugated AuNP (40 nm diameter) and 80 μl of hybridization buffer were added to 500 microtubes. Create a Hybridization Mixture. The mixture is then mixed using a vortex and centrifuged to remove the solution from the lid. Subsequently, the third preliminary biochip is placed in a hybridization chamber filled with water at the bottom. Subsequently, the hybridization mixture is injected while being careful not to generate bubbles through the sample inlet existing in the cover covering the third preliminary biochip. Then close the cover of the hybridization chamber. Subsequently, the mixture is placed in a hybridizer at room temperature and reacted for 1 hour. Next, the cover of the hybridized third preliminary biochip is removed. The washing is then performed by shaking or stirring at 50 rpm for 2 minutes in a washing buffer. Next, washing is performed by shaking or stirring at 50 rpm for 1 minute in distilled water. The third preliminary biochip is then centrifuged at 1000 rpm for 1 minute or dried using compressed air. As a result, a fourth preliminary biochip to which the target DNA indicated by nanoparticles is attached to the probe DNA is prepared.

(5) above GST layer of Phase change  Laser probe for

Specifically, referring to FIG. 8, the GST is crystallized by irradiating a laser after attaching the above-described probe DNA and AuNP-target DNA. In this case, when the target probe DNA having a base sequence complementary to the fixed probe DNA is attached to a high density, the degree of phase change of the GST is reduced because the laser is blocked by the nanoparticles of the target probe is large. something to do. On the other hand, when a target probe DNA having a base sequence complementary to the fixed probe DNA is attached at low density, the degree of phase change of the GST is decreased because the laser is blocked by the nanoparticles of the target probe. Will do a lot. Thereby, the laser-irradiated biochip is completed.

(6) resistance measurement

As described above, the bio information may be detected using a resistance signal detected in the phase change biochip using a system configuration diagram capable of detecting electrical characteristics of the biochip including the phase change layer of FIG. 1.

[Experimental Example]

After fabricating a biochip on which a substrate, an electrode, a phase change material, and a protective film were formed, an arbitrary SPOT was selected before the laser was irradiated to measure resistance, and the resistance of the SPOT was measured after laser irradiation. The measurement results are shown in FIG. 9 and Table 2.

Laser irradiation (with or without) Board electrode Phase change material Measuring resistance radish Si Ti / TiN GST 10 ⁸ Ω U Si Ti / TiN GST 10 ⁵ Ω

1 is a system configuration diagram capable of detecting the resistance of a biochip including a phase change layer.

2 is a LAY_OUT diagram for manufacturing a biochip including a phase change layer.

3 is an enlarged view of a portion A showing a SPOT in which the organic material is formed in FIG. 2.

FIG. 4 is a vertical view illustrating an electrode forming process of a biochip by cutting FIG. 3 into BB ′.

FIG. 5 is a vertical view illustrating a GST / SiO ₂ formation process of a biochip on FIG. 4.

6 is SiO ₂ It is the vertical degree which shows surface treatment.

FIG. 7 is a vertical view schematically illustrating the fixing of target DNA labeled with probe DNA and nanoparticles. FIG.

FIG. 8 is a vertical view schematically illustrating a method of irradiating a laser beam onto a biochip to which target nanoparticles having nanoparticles are attached.

FIG. 9 is a graph showing resistance values before and after laser irradiation to SPOT sites of samples that have been processed up to an electrode, a phase change material, and a protective film forming process.

Claims (10)

In a biochip including a substrate, an electrode, a phase change layer, a protective film, an organic material, and nanoparticles, the phase change layer may include Te, Se, Ge, Sb, Bi, Pb, Sn, As, In _, Ag, S, Si Chalcogenide material consisting of at least one element selected from the group consisting of P, O, The protective film is a biochip, characterized in that the oxide film or nitride film. The method of claim 1, The phase change layer is Ge _a Sb _b Te _c (a, b, c is the atomic mole fraction, a + b + c = 1, 0 <a, b, c <1) or In _d Ag _e Sb _f Te _g ( d, e, f, g are each an atomic mole fraction, d + e + f + g = 1, 0 <d, e, f, g <1). The method of claim 1, The electrode includes titanium (Ti), titanium nitride (TiN), tungsten (tungsten, W), gold (Au), aluminum (Aluminum, Al), tantalum (Ta), tantalum nitride (TaN), A biochip comprising a metal selected from the group consisting of molybdenum (Molybdenum, Mo) and copper (Copper, Cu). delete The method of claim 1, The organic material is a biochip, characterized in that at least one selected from the group consisting of DNA, PNA, proteins and pigments. The method of claim 1, The organic material and the fixed probe fixed to the phase change layer Biochip, characterized in that it comprises a target probe that is complementary to the fixed probe. The method of claim 6, The nanoparticle is a biochip, characterized in that coupled to the target probe. The method of claim 1, The protective film is a biochip, characterized in that the surface treatment. Forming a protective film on a biospot of a phase change layer connected between electrodes formed on the substrate; Preparing a biochip to which a fixed probe which is an organic material is fixed to the protective film; Bonding the fixed probe to the biochip by spraying a target probe having a nanoparticle bound to the biochip; Irradiating laser light onto the biospot; And detecting a phase change by measuring resistance between the electrodes. The phase change layer is characterized by using a chalcogenide material consisting of at least one element selected from the group consisting of Te, Se, Ge, Sb, Bi, Pb, Sn, As, S, Si, P, and O. A detection method using a biochip according to any one of claims 1 to 3 and 5 to 8. A diagnostic kit comprising the biochip of any one of claims 1 to 3 and 5 to 8.

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