CN110639629A - A nano-pillar array microfluidic chip and its detection method - Google Patents

Abstract

The invention discloses a nano-pillar array microfluidic chip and a detection method thereof. The chip comprises a bottom chip, a probe laying layer chip and a sample introduction layer chip which are used in combination with the chip, and a probe laying layer chip is provided on the probe laying layer chip. The needle lays a flow channel, and the sample introduction layer chip is provided with a sample introduction flow channel. When in use, the probe lays a flow channel and the sample introduction flow channel is vertically arranged; m×n detection areas are arranged on the bottom chip , where m is the number of probe-laid flow channels, n is the number of sample-injection channels, and the detection area is located at the intersection of the probe-laid flow channel and the sample-injection channel, and includes an array composed of a large number of nanopillars The chip and the detection method disclosed by the invention have small sample consumption, high detection sensitivity, high detection accuracy, and increased capture efficiency, and are especially suitable for the fluorescence detection of DNA, RNA and protein.

Description

A nano-pillar array microfluidic chip and its detection method

technical field

The invention relates to microfluidic technology, in particular to a nano-pillar array microfluidic chip and a detection method thereof.

Background technique

Fluorescent label detection has a wide range of applications in marine science, biology, and medical research. Using fluorescent labeling detection, it can quantitatively detect DNA, RNA, protein and other organic substances. However, in fluorescence detection, the intensity of fluorescence intensity will directly affect the sensitivity and accuracy of fluorescence detection. Fluorescent biomarkers and specific probes are extremely expensive, and the extraction process of DNA, RNA, protein and other biological substances is extremely expensive. It is cumbersome, time-consuming and expensive for reagents. Therefore, if the amount of samples and reagents used can be reduced, the cost of detection will be greatly reduced. In addition, there are also problems such as low fluorescence intensity in the fluorescence detection of non-biochemical substances, which affects the detection sensitivity and accuracy.

SUMMARY OF THE INVENTION

In order to solve the above technical problems, the present invention provides a nano-pillar array microfluidic chip and a detection method thereof, so as to achieve the purpose of small sample consumption, high detection sensitivity, high detection accuracy and increased capture efficiency.

For achieving the above object, technical scheme of the present invention is as follows:

A nano-pillar array microfluidic chip includes a bottom layer chip, a probe laying layer chip and a sample injection layer chip used in combination with the chip, wherein a probe laying flow channel is provided on the probe laying layer chip, and the sample introduction layer A sampling flow channel is set on the chip, and when in use, the probe laying flow channel and the sampling flow channel are arranged vertically; m×n detection areas are arranged on the bottom chip, wherein m is the length of the probe laying flow channel The number, n is the number of the injection channel, the detection area is located at the intersection of the probe laying channel and the injection channel, and includes an array composed of a large number of nano-pillars.

In the above scheme, the probe laying flow channel is a parallel flow channel with injection ports at both ends, the number of the probe laying flow channel is equal to the probe type, the probe laying flow channel width is greater than the width of the detection area, and the probe laying flow channel is larger than the detection area width. The channel depth is greater than the nanopillar height.

In the above scheme, the sample injection channel is a parallel channel with injection ports at both ends, the number of sample injection channels is equal to the number of samples to be detected, and the direction of the sample injection channel is perpendicular to the direction in which the probes are laid. The width of the injection channel is greater than the lateral width of the detection area, and the depth of the injection channel is greater than the height of the nano-column.

In the above solution, the array composed of the nano-pillars is obtained by nano-imprinting, or by the growth of nano-materials.

A method for detecting a nano-pillar array microfluidic chip, using the above-mentioned nano-pillar array microfluidic chip, first using a probe to lay a layer chip to combine with a bottom chip, and injecting different kinds of probes into the probe laying flow channel , so that the specific probe molecules are laid on the nano-pillars; then the probe laying layer chip is removed, and the sampling layer chip is combined with the bottom chip, so that the sampling flow channel is perpendicular to the probe laying flow channel, and the sampling flow channel is perpendicular to the sampling flow channel. Different samples are injected into the channel, and the samples flow through the detection areas with their respective specific probes; finally, they are detected by a fluorescence microscope or a fluorescence scanner. The intensity analysis results in the type and content of specific molecules.

Through the above technical solutions, the nano-pillar array microfluidic chip provided by the present invention is prepared by micro-nano processing technology, and the highly integrated biological chip is very suitable for the fluorescence detection of DNA, RNA and protein. Compared with the prior art, the present invention has the following beneficial effects:

1. The microfluidic chip designed in the present invention can greatly reduce the consumption of detection samples and detection reagents, thereby greatly reducing the cost of sample processing and the cost of detection reagents.

2. The detection areas of the microfluidic chip designed in the present invention are arranged in a spot-like order, and the fluorescence detection bright spots of the same sample for different probes will be arranged in a row, and the detection effect is clear at a glance.

3. There are a large number of nano-pillar arrays in the detection area, and the neatly arranged nano-pillar arrays have fluorescence enhancement effect.

4. Compared with the flat detection area, the nanopillars will have a large number of specific probes in the lateral direction. During fluorescence scanning, the fluorescent signals will be superimposed in the longitudinal direction, thereby enhancing the fluorescence effect.

5. Compared with the flat detection area, the three-dimensional structure of the nanopillar not only increases the number of probes, but also enhances the contact area between the sample and the detection area, so that more specific molecules with fluorescent labels are combined with the probes, and the detection accuracy is improved. and sensitivity will be improved.

Description of drawings

In order to illustrate the embodiments of the present invention or the technical solutions in the prior art more clearly, the following briefly introduces the accompanying drawings that are required in the description of the embodiments or the prior art.

FIG. 1 is a schematic plan view of an underlying chip disclosed in an embodiment of the present invention;

FIG. 2 is a schematic three-dimensional structure diagram of a detection area disclosed in an embodiment of the present invention;

3 is a schematic three-dimensional structure diagram of a probe laying layer chip disclosed in an embodiment of the present invention;

FIG. 4 is a schematic three-dimensional structure diagram of a sample injection layer chip disclosed in an embodiment of the present invention;

FIG. 5 is a schematic plan view of the bottom chip disclosed in the embodiment of the present invention when the probe laying layer chip is combined;

FIG. 6 is a schematic plan view of the bottom chip disclosed in the embodiment of the present invention when the sample introduction layer chip is combined.

In the figure, 1. Bottom chip; 2. Probe laying layer chip; 3. Sample injection layer chip; 4. Detection area; 5. Nano-pillar; 6. Probe laying flow channel inlet; 7. Probe laying flow channel outlet ; 8. The inlet of the injection channel; 9. The outlet of the injection channel; 10. The probe laying the channel; 11. The injection channel.

Detailed ways

The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings in the embodiments of the present invention.

The invention provides a nano-pillar array microfluidic chip and a detection method thereof. The chip and the detection method have the advantages of small sample consumption and high detection sensitivity.

A nano-pillar 5 array microfluidic chip, comprising a bottom chip 1 and a probe laying layer chip 2 and a sample feeding layer chip 3 used in combination with the chip 2, wherein a probe laying flow channel 10 is provided on the probe laying layer chip 2, The sample layer chip 3 is provided with a sample injection channel 11, and when in use, the probe laying channel 10 and the sample injection channel 11 are arranged vertically.

As shown in FIG. 1 and FIG. 2 , m×n detection areas 4 are provided on the bottom chip 1 , where m is the number of the flow channels 10 laid by the probes, n is the number of the sample injection channels 11 , and the detection area 4 It is located at the intersection of the probe laying channel 10 and the injection channel 11 , and includes an array composed of a large number of nano-pillars 5 .

As shown in FIG. 3, the probe laying flow channel 10 is a parallel flow channel with injection ports at both ends. As shown in FIG. 3, one end of the flow channel is the probe laying flow channel inlet 6, and the other end is the probe laying flow channel outlet. 7. The number of the flow channels 10 laid by the probes is equal to the type of the probes, the width of the flow channels is greater than the width of the detection area 4, and the depth of the flow channels is greater than the height of the nano-pillars 5.

As shown in FIG. 4 , the injection channel 11 is a parallel channel with injection ports at both ends. As shown in FIG. 4 , one end of the channel is the inlet 8 of the injection channel, and the other end is the outlet 9 of the injection channel; The number of sample flow channels 11 is equal to the number of samples to be detected, the flow channel direction is perpendicular to the direction in which the probe lays the flow channel 10 , the flow channel width is greater than the lateral width of the detection area 4 , and the flow channel depth is greater than the height of the nanocolumns 5 .

The array composed of nano-pillars 5 is obtained by nano-imprinting, or obtained by nano-material growth.

In the present invention, the number of 10 flow channels laid by the probe determines the maximum number of probe types, and the number of 11 sample injection channels determines the maximum number of samples detected by a single chip. The two flow channels are perpendicular to each other and jointly determine the detection area 4 of the bottom chip 1 number and topology. On the other hand, the underlying chip 1 has a detection area 4 formed by nano-pillars 5 . The detection area 4 can be of any shape, but the length and width of the detection area 4 are respectively smaller than the widths of the probe laying flow channel 10 and the sample injection flow channel 11 , and are located at the intersection of the two upper flow channels. The functionalized bottom chip 1 is combined with the probe laying layer chip 2 in order to lay different specific probes in the detection area 4 of each column. After the probe is combined with the nano-pillars 5 in the detection area 4 of the underlying chip 1, the probe laying layer chip 2 is removed, and then the underlying chip 1 is combined with the sample injection layer chip 3, so that each row of the detection area 4 is in a different input layer. In the sample flow channel, each sample (pre-fluorescently labeled) will flow through each column of detection areas 4, and then come into contact with different specific probes and have different degrees of specific binding.

The detection method of the nano-pillar 5 array microfluidic chip of the present invention is as follows:

First, the probe laying layer chip 2 is combined with the bottom chip 1. As shown in FIG. 5, different kinds of probes are injected into the probe laying channel 10, so that specific probe molecules are laid on the nanopillars 5; Remove the probe laying layer chip 2, and then combine the sample injection layer chip 3 with the bottom chip 1, as shown in Figure 6, inject different samples into the sample injection channel 11, and the samples flow through the respective specific probes. Finally, it is detected by a fluorescence microscope or a fluorescence scanner, and fluorescent spots with different brightness are formed under the excitation of a specific wavelength of laser light, and then the type and content of specific molecules can be obtained through fluorescence intensity analysis.

The above description of the disclosed embodiments enables any person skilled in the art to make or use the present invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the invention. Thus, the present invention is not intended to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims (5)

1. A nano-column array micro-fluidic chip is characterized by comprising a bottom chip, a probe laying layer chip and a sample injection layer chip, wherein the probe laying layer chip and the sample injection layer chip are respectively combined with the bottom chip for use; the bottom chip is provided with m multiplied by n detection areas, wherein m is the number of probe laying runners, n is the number of sample injection runners, and the detection areas are positioned at the intersection of the probe laying runners and the sample injection runners and comprise arrays formed by a large number of nano-columns.

2. The nanopillar array microfluidic chip according to claim 1, wherein the probe-laying flow channels are parallel flow channels having injection ports at both ends, the number of the probe-laying flow channels is equal to the number of the probe types, the width of the probe-laying flow channels is greater than the width of the detection region, and the depth of the probe-laying flow channels is greater than the height of the nanopillars.

3. The nanopillar array microfluidic chip according to claim 1, wherein the sample injection flow channels are parallel flow channels with injection ports at both ends, the number of the sample injection flow channels is equal to the number of samples to be detected, the direction of the sample injection flow channels is perpendicular to the direction of the probe laying flow channels, the width of the sample injection flow channels is larger than the lateral width of the detection area, and the depth of the sample injection flow channels is larger than the height of the nanopillars.

4. The nanopillar array microfluidic chip according to claim 1, wherein the array of nanopillars is obtained by nanoimprint or by nanomaterial growth.

5. A detection method of a nano-column array micro-fluidic chip adopts the nano-column array micro-fluidic chip as claimed in claim 1, and is characterized in that a probe laying layer chip is firstly used for being combined with a bottom chip, and different types of probes are injected into a probe laying flow channel, so that specific probe molecules are laid on nano-columns; then uncovering the probe laying layer chip, combining the sample introduction layer chip with the bottom layer chip to ensure that the sample introduction flow channel is vertical to the probe laying flow channel, injecting different samples into the sample introduction flow channel, wherein the samples flow through the detection areas respectively provided with the respective specific probes; and finally, detecting by a fluorescence microscope or a fluorescence scanner, forming fluorescent light spots with different brightness under the excitation of laser with specific wavelength, and further obtaining the type and the content of the specific molecules by fluorescence intensity analysis.

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