CN213022964U

CN213022964U - Microarray analysis and detection device

Info

Publication number: CN213022964U
Application number: CN202021798921.4U
Authority: CN
Inventors: 杈逛豢; 边仿
Original assignee: Tianjin Haixing Technology Co ltd
Current assignee: Tianjin Haixing Technology Co ltd
Priority date: 2020-08-25
Filing date: 2020-08-25
Publication date: 2021-04-20
Anticipated expiration: 2030-08-25

Abstract

The utility model provides a microarray sugar metabolic analysis detection device, the device includes: the micro-injection pump comprises a control unit, a signal acquisition unit, a micro-array sensor unit and a micro-injection pump, wherein the control unit is connected with the micro-injection pump and the signal acquisition unit, the micro-array sensor unit is connected with the signal acquisition unit, the micro-array sensor unit comprises a circular electrode array which is distributed by 6 multiplied by 6, and the diameter of the circular electrode is 5-50 mu m; the micro-injection pump is controlled by the control unit to quantitatively add sugar solution to the micro-array sensor unit, the signal acquisition unit acquires an electric signal generated by the micro-array sensor unit stimulated by the sugar solution, the signal acquisition unit transmits data to the control unit, and the control unit records and processes the data. The utility model discloses a microarray carbohydrate metabolic analysis detection device sensitivity is high, stability is good, preparation and easy operation.

Description

Microarray analysis and detection device

Technical Field

The application relates to the field of biosensors, in particular to a microfluidic analysis and detection device.

Background

The taste sense is a sense produced by food in the oral cavity to stimulate the chemical sensing system of taste organ, and includes four basic taste senses of sweet, sour, bitter and salty. The various tastes that people usually taste are the result of the mixture of the four tastes. Taste receptors are taste buds on the tongue, which are oval and mainly composed of taste cells and supporting cells, wherein microvilli at the top of the taste cells extend towards the taste pores and contact with saliva, and nerve fibers are innervated at the cell base. The chemicals in the food dissolve in the saliva and contact the taste receptor cells through the taste pores, where they interact with taste receptors or ion channels. These interactions trigger intracellular signaling cascades that induce cellular action potentials, which are ultimately transmitted to the brain via nerve fibers, forming the sense of taste. Taste cells have a number of taste-sensing molecules on their surface, and different substances can bind to different taste-sensing molecules to present different tastes. The taste of human is stimulated by food to feel the taste only for 1.5-4.0ms, which is faster than the visual sense for 13-45ms, the auditory sense for 1.27-21.5ms and the tactile sense for 2.4-8.9 ms. Conventionally, taste analysis is mainly determined by an artificial taste analysis method, which has a high subjectivity and is difficult to accurately analyze and evaluate taste brought by food. Therefore, the taste sensor can realize on-site, rapid and real-time detection, and has important significance for the food field. Particularly, dynamic detection under the flow state caused by food consumption is more necessary.

Sugar is an important flavoring agent, and is divided into high-calorie sweeteners and low-calorie sweeteners according to the caloric content. The high calorie sweetener comprises sucrose, honey and other natural sweeteners, and has high calorie, and is easy to cause obesity and even diabetes after long-term over-consumption. Low calorie sweetener refers to a material having sweetness, low caloric power and low nutritional value, which is commonly used for controlling blood sugar elevation, preventing obesity, controlling body weight and preventing cardiovascular disease, and is also used as a sugar substitute for diabetics. Lactic acid bacteria in the oral cavity of humans and other mammals ferment with trace amounts of sugars in food to produce organic acids, including lactic acid, acetic acid (ethanol), and the like, which impart specific flavors to various sugars. The metabolic characteristics of the lactic acid bacteria on different saccharides are accurately analyzed through the biosensor, and the method has great significance for preparing the sweetening agent with low calorie and the taste close to sucrose in the field of food.

Disclosure of Invention

The utility model aims at the shortcoming that prior art exists, provide a microarray analysis detection device, solve the conventional taste analysis that exists among the prior art and detect that the subjectivity is strong, the static accuracy subalternation problem that brings of detection thing.

The utility model provides a microarray sugar metabolic analysis detection device, the device includes: the micro-injection pump comprises a control unit, a signal acquisition unit, a micro-array sensor unit and a micro-injection pump, wherein the control unit is connected with the micro-injection pump and the signal acquisition unit, the micro-array sensor unit is connected with the signal acquisition unit, the micro-array sensor unit comprises a circular electrode array which is distributed by 6 multiplied by 6, and the diameter of the circular electrode is 5-50 mu m; the micro-injection pump is controlled by the control unit to quantitatively add sugar solution to the micro-array sensor unit, the signal acquisition unit acquires an electric signal generated by the micro-array sensor unit stimulated by the sugar solution, the signal acquisition unit transmits data to the control unit, and the control unit records and processes the data.

Preferably, the control unit comprises a temperature control module for controlling the detected temperature.

Preferably, the signal acquisition unit comprises a multi-channel amplifier and an analog-to-digital conversion circuit, the multi-channel amplifier is used for amplifying detection signals of different electrodes of the microarray sensor, and the analog-to-digital conversion circuit is used for converting the detection signals into analog signals.

Preferably, the microarray sensor unit comprises a microelectrode array, a detection cavity and a taste activity layer;

preferably, the detection chamber is affixed to the array of microelectrodes, and the taste-active layer is located within the detection chamber and is affixed to the upper surface of the array of microelectrodes. The microelectrode array comprises an electrode array with regular polygon nodes arranged at equal intervals. The electrode spacing of the microelectrode array is adapted to the taste activity unit density.

Preferably, the taste-active layer comprises biologically active taste cells. The taste-active layer is attached to the upper surface of the microelectrode array with the taste pores of the taste cells facing upward. And arranging a tensioned and stretched PMMA net on the taste activity layer to enable the taste activity layer to be tightly attached to the microelectrode array.

Preferably, a layer of polyornithine and laminin is formed between the taste-active layer and the microelectrode array.

Compared with the prior art, the utility model discloses the beneficial effect of embodiment is: the utility model obtains the micro-array sensor with high sensitivity and high stability by organically combining the microelectrode array and the gustatory active layer, and simplifies the preparation and operation of the gustatory sensor; the taste cells are fixed through the PMMA net, the polyornithine and the laminin layer, so that the taste active layer is effectively fixed, and the detection stability of the sensor is improved; based on the utility model discloses a sugar lactic acid device carries out sugar taste simulation and detects, the accurate acidity scope that obtains the compound sweetener that metabolic taste and cane sugar are close.

Drawings

The above features and advantages of the present invention will become more apparent and readily appreciated from the following description of the exemplary embodiments thereof taken in conjunction with the accompanying drawings.

FIG. 1 is a schematic view showing the structure of a microarray carbohydrate metabolism analysis/detection apparatus according to the present invention.

Fig. 2 is a schematic structural view of a microarray sensor unit according to the present invention.

Detailed Description

The present invention will be described in further detail with reference to the attached drawings so that those skilled in the art can accurately understand the present invention.

As shown in fig. 1, the microarray carbohydrate metabolism analysis and detection apparatus 100 provided in this embodiment includes: a control unit 110, a signal acquisition unit 120, a microarray sensor unit 130, and a micro syringe pump 140. The micro syringe pump 140 is connected to the control unit 110, the control unit 110 controls the quantitative addition of the sugar solution for detection to the microarray sensor unit 130, and the microarray sensor unit 130 is connected to the signal acquisition unit 120 to realize the signal acquisition of the microarray sensor unit 130. The signal acquisition unit 120 is connected to the control unit 110 via a serial port to complete data transmission, recording and processing.

The micro syringe pump 140 may control the injection amount of the test solution through an asynchronous receive/transmit protocol (UART). The signal collection unit 120 includes a multi-channel amplifier for amplifying voltage signals of each channel of the micro-array sensor unit 130, and the signal collection unit 120 further includes an analog-to-digital conversion point for converting the detected voltage signals into digital signals and transmitting the digital signals to the control unit 110. The microarray carbohydrate metabolism analysis and detection apparatus 100 further includes a control unit 110 including a temperature control module for controlling the temperature of the analysis and detection, typically at 37 ℃ to 38 ℃.

[ microarray sensor Unit ]

As shown in FIG. 2, the microarray sensor unit 130 of the present embodiment includes a micro-electrode array 131, a detection chamber 132, and a taste-active layer 133. Microelectrode array 131 comprises an array of regularly-spaced polygonal-node electrodes to avoid excessive electrodes leading to increased interference, difficult lead placement, and reduced complexity of parallel analysis. The electrode spacing is adapted to the taste activity cell density to reduce interference of adjacent taste activity cells. The detection cavity 132 is fixed on the microelectrode array 131, and the taste activity layer 133 is positioned in the detection cavity 132 and attached to the upper surface of the microelectrode array 131. The process flow for preparing the microarray sensor unit 130 includes:

step S110: selecting a silicon substrate with the thickness of 450 mu m, cleaning the silicon substrate by using acetone, heating the silicon substrate at 200 ℃ for 30min, drying the silicon substrate, and cutting the silicon substrate into a silicon wafer with the flat length of 6mm by using laser;

step S120: coating a photoresist film (such as negative photoresist) with a thickness of 20-150 μm on a silicon wafer, drying by a step method, keeping at 65 ℃ for 1 minute, heating to 95 ℃ and repelling for 5-10 minutes, cooling to 65 ℃ and keeping for 1 minute, and then standing and cooling;

step S130: the adopted wavelength is 365nm, and the light intensity is 13mw/cm²Exposing for 5-30 seconds, wherein the pattern on the mask is a circular hole array distributed by 6 multiplied by 6, the diameter of the circular holes is 5-50 mu m, the distance between the circular holes is 1-2 times of the diameter, and the circular holes are through holes;

step S140: keeping at 65 deg.C for 1 min, heating to 95 deg.C, repelling for 5-10 min, standing, cooling, developing with developer PGMEA for 5 s-2 min, and curing at 150 deg.C for 5-10 min;

step S150: filling the round holes on the silicon chip with the round hole array by using a vacuum evaporation method, a vacuum sputtering method, a vacuum ion plating method or a chemical vapor deposition method to form a conducting layer with the thickness of 20 nm-80 nm, wherein the conducting layer is selected from any one of gold, silver, copper, nickel, titanium, platinum and the like;

step S160: forming an electrode array and a guide line of each electrode by dry etching to form an electrode array chip, wherein the electrode array chip comprises a circular electrode array distributed by 6 multiplied by 6, and the diameter of the circular electrode is 5-50 mu m;

step S170: fixing a reference detection cavity on the surface of the electrode array chip by using the diameter of glue with biological stability, wherein the side length of the detection cavity is a PMMA frame with 5 mm;

step S180: the gustatory activity tissue layer is placed into the detection cavity and attached to the upper surface of the electrode array chip in a mode that the taste holes face upwards, gustatory nerves are communicated with the electrodes, and the tissue layer is tightly attached to the electrode array through the PMMA net stretched and tensioned on the gustatory activity tissue layer, so that the electrical connection is promoted.

To facilitate the attachment of the taste-active layer 133 to the microelectrode array 131, the device was immersed in a phosphate buffer containing 0.1mg/ml polyornithine for 24 hours at room temperature and then in a phosphate buffer containing 8g/ml laminin for 24 hours to form a 4nm layer of polyornithine and laminin between the taste-active layer and the microelectrode array, which promotes the adhesion of the taste-active layer to the microelectrode array due to the positive charge of polyornithine and laminin.

[ taste-active layer ]

In the taste-active layer of this example, SD rats weighing about 250g were selected, which had intact taste cells obtained by isolating the tongue epithelial tissue of mice. The preparation process comprises the following steps:

step S210: fixing SD rat, injecting urethane on abdomen for anesthesia;

step S220: cutting the tongue to the near end of the annular papilla, peeling off the fungiform papilla of the tongue, hatching with ringer's solution and rinsing for three times;

ringer's solution was prepared by adding 8.6g of sodium chloride, 0.3g of potassium chloride and 0.28g of calcium chloride to 1000ml of distilled water.

Step S230: injecting 0.4ml injection buffer solution to the upper and lower skin of tongue;

the buffer contained 1.5mg/ml collagenase type II (Gibco) and 3mg/ml racemase type II (Roche).

Step S240: incubating with ringer's solution at room temperature for 8-10 min, peeling tongue epithelium from lower muscular layer, and adding into ringer's solution;

step S250: the tongue epithelium was separated into 5mm by 5mm format and rinsed with ringer's solution to give a taste active layer.

The present invention is described in detail with reference to the embodiments, but it can be understood by those skilled in the art that the above embodiments are only one of the preferred embodiments of the present invention, and for space limitation, all embodiments can not be listed herein, and any implementation that can embody the technical solution of the claims of the present invention is within the protection scope of the present invention.

It should be noted that the above is a detailed description of the present invention, and it should not be considered that the present invention is limited to the specific embodiments, and those skilled in the art can make various modifications and variations on the above embodiments without departing from the scope of the present invention.

Claims

1. A microarray carbohydrate metabolism analysis and detection device, comprising: the micro-injection pump comprises a control unit, a signal acquisition unit, a micro-array sensor unit and a micro-injection pump, wherein the control unit is connected with the micro-injection pump and the signal acquisition unit, the micro-array sensor unit is connected with the signal acquisition unit, the micro-array sensor unit comprises a circular electrode array distributed by 6 x 6, and the diameter of the circular electrode is 5-50 mu m;

the micro-injection pump is controlled by the control unit to quantitatively add sugar solution to the micro-array sensor unit, the signal acquisition unit acquires an electric signal generated by the micro-array sensor unit stimulated by the sugar solution, the signal acquisition unit transmits data to the control unit, and the control unit records and processes the data.

2. The microarray carbohydrate metabolism analysis and detection device according to claim 1, characterized in that: the control unit comprises a temperature control module for controlling the detection temperature.

3. The microarray carbohydrate metabolism analysis and detection device according to claim 1, characterized in that: the signal acquisition unit comprises a multi-channel amplifier and an analog-to-digital conversion circuit, the multi-channel amplifier is used for amplifying detection signals of different electrodes of the micro-array sensor, and the analog-to-digital conversion circuit is used for converting the detection signals into analog signals.

4. The microarray carbohydrate metabolism analysis and detection device according to claim 1, characterized in that: the microarray sensor unit comprises a microelectrode array, a detection cavity and a taste active layer;

the detection cavity is fixed on the microelectrode array, and the gustatory activity layer is positioned in the detection cavity and is attached to the upper surface of the microelectrode array.

5. The microarray carbohydrate metabolism analysis and detection device according to claim 4, characterized in that: the microelectrode array comprises an electrode array with regular polygon nodes arranged at equal intervals.

6. The microarray carbohydrate metabolism analysis and detection device according to claim 5, characterized in that: the electrode spacing of the microelectrode array is adapted to the taste activity unit density.

7. The microarray carbohydrate metabolism analysis and detection device according to claim 4, characterized in that: the taste-active layer includes biologically active taste cells.

8. The microarray carbohydrate metabolism analysis and detection device according to claim 7, characterized in that: the taste active layer is attached to the upper surface of the array of microelectrodes with the taste pores of the taste cells facing upwards.

9. The microarray carbohydrate metabolism analysis and detection device according to claim 8, characterized in that: and arranging a tensioned and stretched PMMA net on the taste activity layer to enable the taste activity layer to be tightly attached to the microelectrode array.

10. The microarray carbohydrate metabolism analysis and detection device according to claim 8, characterized in that: a layer of polyornithine and laminin is formed between the taste-active layer and the microelectrode array.