CN103336112A - Method for detecting human immunoglobulin E by adopting carbon nano tube micro-cantilever biosensor - Google Patents

Abstract

The invention discloses a high-sensitivity, fast and accurate detection method capable of rapidly measuring human immunoglobulin E (hIgE), which is realized by constructing a carbon nanotube micro-cantilever biosensor. The biosensor includes a support, a base material, a carbon nanotube, a pick-up circuit, and a layer of nucleic acid aptamer is modified on the carbon nanotube. Firstly, a detection probe containing hIgE nucleic acid aptamer is made on the carbon nanotube micro-cantilever. During detection, the detection probe is put into the sample to be tested, and the hIgE in the sample to be tested reacts specifically with the nucleic acid on the detection probe. The aptamer forms a complex and attaches to the micro-cantilever; the mass change of the complex on the micro-cantilever causes the flexural displacement or the change of the resonance frequency of the micro-cantilever and the relationship between the mass of the complex and the sample to be tested The concentration of hIgE is positively correlated, so as to realize the detection of hIgE.

Description

Method for detecting human immunoglobulin E with carbon nanotube microcantilever biosensor

technical field

The invention relates to the field of biomedical engineering, in particular to a method for detecting hIgE with a micro-cantilever beam biosensor.

technical background

Human immunoglobulin E (Human immunoglobulin E, h IgE) is a common tumor marker. Currently, there are many methods for detecting h IgE, including radioimmunoassay, enzyme-linked immunoassay, and fluorescent immunoassay. Among them, the radioimmunoassay method has radiation and is not safe; other detection methods are complicated to operate and have low sensitivity, making it difficult to realize the diagnosis and research of early diseases, and cannot meet the needs of rapid detection. It is necessary to establish a rapid, sensitive and easy-to-operate hIgE detection method.

Contents of the invention

The technical problem to be solved by the present invention is to provide a detection method with high sensitivity, simple structure, fast and accurate, which can realize rapid determination of hIgE.

In order to solve the above technical problems, the present invention realizes the detection of hIgE by constructing a carbon nanotube microcantilever biosensor. The carbon nanotube micro-cantilever biosensor includes a support, a base material, carbon nanotubes, and a pick-up circuit; wherein the base material is fixed on one side of the support to form a micro-cantilever structure, carbon nanotubes are grown on the base material, and the pick-up circuit is on the base Below the material, a layer of nucleic acid aptamer is also modified on the carbon nanotube.

The preparation steps of the carbon nanotube microcantilever biosensor used in the present invention are as follows:

1. Fabrication of micro-cantilever structure

The micro-cantilever beam is processed into a micro-cantilever beam structure by using the semiconductor material silicon as the base material.

2. Production of pick-up circuit

The pick-up circuit is to manufacture silicon varistors by using microelectronic technology on the lower surface of the base material, and connect four varistors to form a Wheatstone bridge.

3. Cantilever beam growth and carbon nanotube coating process

The upper surface of the base material in the preceding steps is cleaned by ultrasonic cleaning with acetone, absolute ethanol, and deionized water respectively, and then carbon nanotubes are grown by low-pressure chemical vapor deposition (LPCVD). Carbon nanotubes can also be coated on silicon substrates by pyrolysis, coating, or other methods.

4. Modification of nucleic acid aptamers on carbon nanotube microcantilevers

The nucleic acid aptamer is modified on the carbon nanotube through hydrophobic interaction to form a detection probe that can specifically recognize hIgE, thereby constructing the carbon nanotube micro-cantilever biosensor; the hIgE nucleic acid aptamer can also be obtained through π-π stacking effect on carbon nanotubes.

The steps that the present invention detects to hIgE are as follows:

(1) Fabricate a detection probe containing hIgE nucleic acid aptamer on the carbon nanotube microcantilever;

(2) Put the detection probe into the sample to be tested, and the hIgE in the sample to be tested forms a complex with the nucleic acid aptamer on the detection probe through a specific reaction and attaches to the micro-cantilever beam.

(3) The mass of the formed complex is positively correlated with the concentration of hIgE in the sample to be tested.

(4) The mass change of the compound on the micro-cantilever causes the flexural displacement of the micro-cantilever or the change of the resonance frequency, thereby realizing the detection of h IgE.

Description of drawings

Figure 1 is a schematic diagram of a carbon nanotube microcantilever biosensor for detecting hIgE.

Detailed ways

1 is a schematic diagram of a carbon nanotube microcantilever biosensor for h IgE detection, including a support 1 , a base material 2 , a carbon nanotube 3 and a pick-up circuit 4 . Wherein the base material 2 is fixed on one side of the support 1 to form a micro-cantilever beam structure, the carbon nanotube 3 is grown on the base material 2, the pick-up circuit 4 is under the base material 2, and a layer of nucleic acid adapter is also modified on the carbon nanotube 3. Ligand 5.

First, modify the nucleic acid aptamer 5 that specifically recognizes hIgE on the carbon nanotube 3 to form a detection probe; the nucleic acid aptamer 5 can be modified on the carbon nanotube 3 by hydrophobic interaction, or can be Modified on carbon nanotubes 3 by π-π superposition.

Then, the detection probe is put into the sample to be tested, and the nucleic acid aptamer 5 in the probe has a specific recognition reaction with the hIgE in the sample to form a complex, which generates mass on the micro-cantilever biosensor. Effect, using this mass effect to realize the detection of hIgE.

Example 1

The step that the present invention detects hIgE is as follows:

(1) Place the carbon nanotube micro-cantilever in a solution containing a nucleic acid aptamer that has a specific recognition effect on hIgE, and modify the nucleic acid aptamer on the carbon nanotube by ultrasonic treatment. The method can be that the nucleic acid aptamer is modified on the carbon nanotube through hydrophobic interaction, or can be modified on the carbon nanotube through the π-π superposition effect to form a detection probe comprising the hIgE nucleic acid aptamer;

(2) Drop the sample to be tested onto the carbon nanotube microcantilever modified with the nucleic acid aptamer, and incubate at room temperature for 15 minutes, so that the nucleic acid aptamer on the biosensing interface and the h in the sample to be tested IgE undergoes a specific recognition reaction and forms a complex;

(3) The mass of the formed complex is positively correlated with the concentration of hIgE in the sample to be tested.

(4) The formed complex produces a mass effect on the micro-cantilever, which is used to detect hIgE.

In this experiment, 10 μg/ml, 1 μg/ml, and 0.5 μg/ml were sampled, and the mass effects of the formed complexes on the silicon micro-cantilever were 47 Hz, 4.2 Hz, and 2.3 Hz, respectively, and the detection results of h IgE were 10.4 μg /ml, 0.93 μg/ml, 0.51 μg/ml.

Claims (4)

1. detect the method for human immunoglobulin(HIg) E with carbon nano-tube micro-cantilever biology sensor, it detects to adopt carbon nano-tube micro-cantilever biology sensor, it is characterized in that: comprise the steps

(1) on the carbon nano-tube micro-cantilever, makes the detector probe that contains h IgE aptamer earlier;

(2) detector probe is put into sample to be tested, hIgE forms compound by the aptamer on specific reaction and the detector probe and is attached on the micro-cantilever in the sample to be tested;

(3) concentration of h IgE is proportionate in the quality of formed compound size and the sample to be tested;

(4) mass change that produces at micro-cantilever of described compound causes micro-cantilever deflection displacement or change of resonance frequency, thereby realizes the detection to h IgE.

2. according to the said method of claim 1, described carbon nano-tube micro-cantilever biology sensor comprises support (1), base material (2), carbon nano-tube (3), pick-up circuit (4); Described base material (2) is fixed on support (1) one side and constitutes micro cantilever structure, carbon nano-tube (3) be grown in base material (2) above, pick-up circuit (4) is below base material (2); It is characterized in that: on carbon nano-tube (3), also be modified with one deck aptamer (5); The method of described modification is that aptamer passes through hydrophobic effect.

3. according to the said method of claim 2, it is characterized in that: the method for described modification is by π-π superposition.

4. according to the said method of claim 2, it is characterized in that: the method for described growth is Low Pressure Chemical Vapor Deposition.

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