TWI458975B - Immediate electrochemical detection of wafers - Google Patents

Description

Instant electrochemical detection wafer

This invention relates to a wafer, and more particularly to an instant electrochemical detection wafer.

With the rapid development of semiconductor process technology and MEMS technology, microchips fabricated using MEMS technology are a novel and promising industry. Biochips are one of the most promising future star industries. Biochips are micro-devices that use semiconductors for bioanalysis. Bio-wafers are usually based on germanium wafers, glass, or polymers. Structures or control circuits are fabricated to detect and analyze biological molecules or further applications.

The most common method currently used to detect DNA is to perform DNA polymerase chain reaction (PCR). PCR is carried out in three steps that are repeated in sequence, denaturation (denature, DNA double-strand separation to form a single strand) → adhesion (annealing, primer binding to a complementary position on a single strand of DNA) → extension (extension, with two primers A new DNA was synthesized as a starting point. After the PCR was completed, gel electrophoresis was performed to detect the result.

Generally, the number of cycles of PCR is 20 to 30 times, and gel electrophoresis needs to wait until the PCR is completely finished. It is not only time-consuming, but also requires plastics and many consumables to be purchased, and can be read with large optical instruments. The rubber powder is quite expensive, so the use of gel electrophoresis is time consuming and costly.

If gel electrophoresis is not used, capillary electrophoresis can also be used. After the end of PCR, the PCR product is placed in a capillary tube for separation. Although this method does not require glue, it is necessary to purchase consumables such as capillaries and cooperate with large optical instruments. To be able to interpret, it is also time consuming and costly.

In addition, if other electrochemical detection methods are to be used, it is necessary to wait until the end of the PCR and obtain the PCR product, and then add a specific exonuclease for cutting, and the exo-enzyme is also quite expensive and requires a long working time. Not ideal for use.

Accordingly, it is an object of the present invention to provide an instant electrochemical detection wafer that can instantly detect detection results.

Therefore, the instant electrochemical detection wafer of the present invention comprises a reaction device, a first flow rate control device connected to the reaction device, a heating device disposed under the reaction device, and a detection device disposed above the reaction device. Device.

The reaction device includes a substrate, and a flow path formed on the substrate and repeatedly bent and extended. The flow prop has an inlet and an outlet opposite to the inlet.

The first flow rate control device includes a first flow rate controller, and a first injector mounted to the first flow rate controller and communicating with the flow path, the first syringe containing the sample and the reagent.

The heating device includes a first heating unit, a second heating unit, and a third heating unit located below the substrate and spaced apart from each other, and the flow path is repeated across the first to third heating units.

The detecting device includes a cover correspondingly disposed on the substrate, a plurality of electrodes disposed on the cover and extending in the flow channel, and a plurality of probes respectively disposed on the electrodes, and A current detector electrically coupled to the electrode.

The effect of the present invention is that when the sample and the reaction reagent are injected into the flow channel by the first syringe for PCR, the single-stranded DNA formed by denaturation in the sample can be directly bonded to the probe on the electrode, and the change is made. The value of the current detected by the electrode, whereby the result of the PCR can be obtained quickly and directly during the course of PCR.

The above and other technical contents, features and advantages of the present invention will be apparent from the following detailed description of the preferred embodiments.

Referring to Figures 1 and 2, a preferred embodiment of the instant electrochemical detection wafer of the present invention comprises a reaction device 2, a first flow rate control device 3 connected to the reaction device 2 (see Fig. 2), and a The heating device 4 under the reaction device 2, the detecting device 5 disposed above the reaction device 2, and the clamping device 6 for clamping the reaction device 2, the heating device 4, and the detecting device 5 And a second flow rate control device 7 (see FIG. 2) connected to the heating device 4, and the clamping plate 6 is omitted in FIG. 2 for convenience of explanation.

The reaction device 2 includes a substrate 21, and a flow channel 22 formed on the substrate 21 and recessed and extended. The flow channel 22 has an inlet 221 and an outlet 222 opposite to the inlet 221.

As shown in FIG. 1 , the heating device 4 includes a heat transfer plate 41 located below the substrate 21 of the reaction device 2 and corresponding to the size of the substrate 21 , and a first heating unit 42 located below the substrate 21 and spaced apart from each other. a second heating unit 43, and a third heating unit 44, the heat transfer plate 41 has an upper surface 411, a lower surface 412, and a water flow channel 413 formed by the lower surface 412, and the lower portion is sandwiched The plate 6 cooperates with the heat transfer plate 41 to form the water flow passage 413 in a closed state.

The first heating unit 42 has a first heater 421 disposed on the lower surface 412 of the heat transfer plate 41, and a first heat conducting member disposed on the upper surface 411 of the heat transfer plate 41 and corresponding to the first heater 421. 422. The second heating unit 43 has a second heat conducting member 431 disposed on the upper surface 411 of the heat transfer plate 41 and corresponding to the water flow passage 413. The third heating unit 44 has a lower surface disposed on the heat transfer plate 41. a second heater 441 of 412, and a third heat conducting member 442 disposed on the upper surface 411 of the heat transfer plate 41 and corresponding to the second heater 441, wherein the water flow channel 413 is located at the first and second heaters 421 Between 441. The flow path 22 is repeatedly bent and heated across the first, second, and third heat conducting members 422, 431, and 442.

The detecting device 5 includes a cover 51 correspondingly disposed on the substrate 21, and a plurality of electrodes 52 disposed on the cover 51 and extending in the flow channel 22, and a plurality of electrodes 52 are respectively disposed on the electrodes The probe on 52 (the probe is not visible to the naked eye, not shown), and a current detector 53 electrically connected to the electrode 52.

As shown in FIG. 2, the first flow rate control device 3 includes a first flow rate controller 31, and a first injector 32 mounted on the first flow rate controller 31 and communicating with the flow path 22, the first The syringe 32 contains a sample and a reagent. The second flow rate control device 7 includes a second flow rate controller 71, and a second injector 72 mounted to the second flow rate controller 71 and containing liquid and communicating with the water flow channel 413 (see FIG. 1). .

In this embodiment, the first heater 421 and the third heater are respectively controlled such that the temperature of the first heat conducting member 422 is 95 ° C, the temperature of the third heat conducting member 442 is 72 ° C, and the second is controlled. The flow rate controller 71 causes the liquid in the second syringe 72 to flow into the water flow channel 413 at a constant flow rate, so that the heat generated by the first and second heaters 422, 442 is absorbed by the liquid in the water flow channel 413. The temperature rose to 55 °C. It should be noted that the above temperature is only the experimental condition set in this embodiment, and the temperature of the first and second heaters 422 and 442 may be changed according to various experiments to adjust the first heat conducting member 422 and the third heat conducting. The temperature of the member 442, and the temperature of the second heat conducting member 431, is controlled by changing the flow rate of the liquid. The faster the flow rate, the lower the temperature, the slower the flow rate, and the higher the temperature.

In the following, the usage of the present invention will be described in conjunction with the experiment. In this embodiment, the temperatures of the first heat conducting member 422, the second heat conducting member 431, and the third heat conducting member 442 are 95 ° C, 55 ° C, and 72 ° C, respectively. And the flow channel 22 is repeatedly bent and repeated across the first, second, and third heat conducting members 422, 431, 442, thereby dividing the flow channel 22 into a plurality of first heat conducting members 422. A denaturation zone 223 (denaturation), a plurality of bonding regions 224 on the second heat conducting member 431, and a plurality of extensions 225 on the third heat conducting member 442. The inlet 221 of the channel 22 is Located in the first denaturation zone 223, and the outlet 222 is located in the last extension zone 225, the sample passes through the denaturation zone 223, the adhesion zone 224, and the extension zone 225 is a cycle of polymerase chain reaction (PCR). .

First, a fragment complementary to the PCR product is confirmed, and the fragment is immobilized on the electrode 52 as a probe, and it is confirmed that the electrode 52 is to be placed at the position of the first few cycles of the PCR, in this embodiment. The electrodes 52 are respectively disposed between the denaturation region 223 and the adhesion region 224 of the fifth cycle, the twelfth cycle, and the eighteenth cycle. Since the manner of providing the probe is easily implemented by those skilled in the art, It is to be noted that the position of the electrode 52 may be changed depending on the experimental conditions, and is not limited to the content disclosed in the embodiment.

Next, the sample to be subjected to PCR and the reaction reagent are mixed to form a reactant, which is placed in the first syringe 32, and the first flow rate controller 31 is activated, and the reactant is injected from the inlet 221 of the flow channel 22. The reactant is then moved within the flow channel 22 at a set rate. The time ratio of the reactants in the denaturation zone 223, the adhesion zone 224, and the extension zone 225 generally approaches 1:1:2, so that the area of the substrate 21 can be increased in design to extend the flow channel 22. The length of the 225 is longer, and the extension of the flow channel 22 may be more curved and the path of the extension zone 225 may be longer. Both of the above may be implemented, and are not limited thereto. The speed control of the first flow rate controller 31 allows the reactants to advance from the extension zone 225 to the next at a faster rate after each reaction of the denaturation zone 223→adhesion zone 224→extension zone 225 is completed. The denatured zone 223 is such that the reactants undergo a cycle of the next denatured zone 223→adhesive zone 224→extension zone 225.

Through the above design, when the PCR product flows through the denaturation zone 223 to decompose the double-stranded DNA into a single-stranded DNA, since the electrode 52 is disposed between the denatured region 223 and the bonding region 224, if the single-stranded DNA has The fragment complementary to the probe will bind to the probe. As the number of single strands of DNA bound to the probe increases, the amount of current detected by the current detector 53 corresponding to the electrode 52 will be lower.

Thereby, not only the PCR product can be detected in the process of PCR reaction, and the PCR time can be saved without waiting for the completion of the PCR reaction; in addition, the interpretation of the detection result directly by the current intensity is not required. Large or expensive consumables, without the need for additional exozymes, can reduce the cost of the experiment.

In summary, the single-stranded DNA in the in vivo can be directly coupled to the probe on the electrode 52 during the PCR process, and the current value detected by the electrode 52 can be changed, thereby being quickly and directly obtained. As a result of the PCR, it is not necessary to re-rubber, purchase a large amount of expensive or expensive consumables, and it is not necessary to cooperate with a large optical instrument to interpret, and the experiment cost can be saved, so that the object of the present invention can be achieved.

The above is only the preferred embodiment of the present invention, and the scope of the invention is not limited thereto, that is, the simple equivalent changes and modifications made by the scope of the invention and the description of the invention are All remain within the scope of the invention patent.

2. . . Reaction device

twenty one. . . Substrate

twenty two. . . Runner

221. . . Entrance

222. . . Export

223. . . Denatured zone

224. . . Bonding zone

225. . . Extended area

3. . . First flow rate control device

31. . . First flow controller

32. . . First syringe

4. . . heating equipment

41. . . Heat transfer plate

411. . . Upper surface

412. . . lower surface

413. . . Water flow channel

42. . . First heating unit

421. . . First heater

422. . . First heat conducting member

43. . . Second heating unit

431. . . Second heat conducting member

44. . . Third heating unit

441. . . Second heater

442. . . Third heat conducting member

5. . . Detection device

51. . . Cover

52. . . electrode

53. . . Current detector

6. . . Clip board

7. . . Second flow rate control device

71. . . Second flow controller

72. . . Second syringe

1 is an exploded perspective view showing a preferred embodiment of the instant electrochemical detection wafer of the present invention;

Fig. 2 is a plan view, and Fig. 1 is additionally explained, and for convenience of explanation, some of the members are omitted in the drawings.

2. . . Reaction device

twenty one. . . Substrate

twenty two. . . Runner

221. . . Entrance

222. . . Export

4. . . heating equipment

41. . . Heat transfer plate

411. . . Upper surface

412. . . lower surface

413. . . Water flow channel

42. . . First heating unit

421. . . First heater

422. . . First heat conducting member

43. . . Second heating unit

431. . . Second heat conducting member

44. . . Third heating unit

441. . . Second heater

442. . . Third heat conducting member

5. . . Detection device

51. . . Cover

52. . . electrode

53. . . Current detector

6. . . Clip board

Claims (3)

An instant electrochemical detection wafer comprising: a reaction device comprising a substrate, and a flow channel formed on the substrate and repeatedly bent and extended, the flow item having an inlet and an outlet opposite to the inlet; a first flow rate control device includes a first flow rate controller, and a first injector mounted on the first flow rate controller and communicating with the flow path, the first syringe containing the sample and the reagent; The heating device comprises a heat transfer plate located below the substrate and corresponding to the size of the substrate, and a first heating unit, a second heating unit and a third heating unit located below the substrate and spaced apart from each other, the flow channel Repeatingly spanning the first to third heating units, wherein the heat transfer plate has an upper surface, a lower surface, and a water flow passage formed by the lower surface recessed, the first heating unit having a setting a first heater on a lower surface of the heat transfer plate, and a first heat conduction member disposed on the surface of the heat transfer plate and corresponding to the first heater, the second heating unit having a heat transfer plate disposed thereon a second heat conducting member having a surface and corresponding to the water flow passage, the third heating unit having a second heater disposed on a lower surface of the heat transfer plate, and a second heat heater disposed on the heat transfer plate surface and corresponding to the second heater a third heat conducting member, wherein the water flow channel is located between the first and second heaters; a detecting device comprising a cover correspondingly disposed on the substrate, and a plurality of the cover members are disposed on the cover and extending An electrode disposed in the flow channel, a plurality of probes respectively disposed on the electrode, and an electrical connection to the electrode a pole current detector; two clamping plates for clamping the reaction device, the heating device, and the detecting device, and one of the clamping plates cooperates with the heat transfer plate to form a closed state of the water flow channel And a second flow rate control device comprising a second flow rate controller, and a second injector mounted to the second flow rate controller and containing the liquid and communicating with the water flow channel. The instant electrochemical detection wafer according to claim 1, wherein the flow path is repeatedly bent and overlaps across the first to third heat conducting members, thereby dividing the flow path into a plurality of a denaturation zone on the first heat conducting member, a plurality of bonding regions on the second heat conducting member, and a plurality of extending regions on the third heat conducting member, the inlet of the flow channel is located in the first denaturation zone, and the outlet It is located in the last extension area. The instant electrochemical detection wafer according to claim 2, wherein the electrode is located between the denaturation zone and the adhesion zone.