CN105925476B - Portable isothermal nucleic acid analyzer based on mobile phone detection - Google Patents

Abstract

The invention discloses a portable isothermal nucleic acid analysis device based on mobile phone detection. The lower isothermal heating unit is arranged at the bottom of the reaction reagent and heats the reaction reagent. The upper layer constant temperature unit is arranged at the top of the reaction reagent and heats the reaction reagent. The heat capacity adjustment unit is arranged Between the lower isothermal heating unit and the upper constant temperature unit, and keep it in close contact with the lower isothermal heating unit and the upper constant temperature unit; the working angle mechanical adjustment unit includes the optocoupler and stepping motor at the bottom, and finds the positioning origin through the optocoupler. The stepper motor drives the lower isothermal heating unit, the reaction reagent, the upper constant temperature unit and the heat capacity adjustment unit to move. The mobile phone is located on the side of the reaction reagent for fluorescence detection. The invention can effectively simplify the complexity of the nucleic acid analysis device, reduce the nucleic acid analysis time, reduce the cost of the nucleic acid analysis device, and realize convenient, mobile, and rapid on-site molecular diagnosis.

Description

Portable isothermal nucleic acid analysis device based on mobile phone detection

technical field

The invention relates to the field of biomedical detection, in particular to a fast and convenient molecular diagnosis device.

Background technique

Compared with other technologies, such as immunoassay technology, molecular diagnostic technology has significant advantages in sensitivity. For example, in HIV detection, immunological detection technology has a window period problem, while molecular diagnostic technology can achieve accurate diagnosis at the earliest stage of HIV infection. Therefore, in recent years, molecular diagnosis has been an important development direction of in vitro diagnostic technology. Molecular diagnostic technology is playing an increasingly important role in ensuring people's health and safety, such as rapid detection and emergency prevention of various highly sudden and highly transmissible diseases such as SARS, avian influenza, and H1N1 influenza In the control, molecular diagnostic technology has played an irreplaceable role.

A typical representative of molecular diagnostic technology is PCR gene amplification technology. With the help of specific thermal cycle technology, PCR gene amplification can perform millions of times of amplification and replication on the original trace nucleic acid template, thereby realizing quantitative or qualitative detection. Two significant limitations of traditional PCR amplification technology are: complex thermal cycle device and long amplification reaction time. Therefore, so far, PCR testing is often performed in central laboratories, which is not conducive to the expansion of the application of PCR technology, such as molecular diagnosis in simple medical institutions, remote areas, and field sites with relatively simple medical facilities. Therefore, the study of new PCR amplification technology, such as rapid type and isothermal amplification PCR technology based on single-point constant temperature heating or two-point constant temperature heating, can simplify the heating device, shorten the detection time, reduce the cost of the device, and realize the improvement of the medical environment. Low dependence, on-site rapid molecular diagnosis has important practical significance. On the other hand, a simplified fluorescence detection technology is used, such as the camera of the mobile phone to detect the fluorescence image signal in the nucleic acid analysis reaction, thus eliminating the need for a dedicated fluorescence detection module. At the same time, the mobile phone can also be used as an intelligent instrument operation terminal to run complex digital image processing algorithms and provide a friendly human-machine interface, which is not only conducive to reducing the cost of molecular diagnosis, but also conducive to the realization of mobile and convenient medical treatment.

Contents of the invention

The invention provides a portable isothermal nucleic acid analysis device based on mobile phone detection, which can effectively simplify the complexity of the nucleic acid analysis device, reduce the nucleic acid analysis time, reduce the cost of the nucleic acid analysis device, and realize convenient, mobile, and on-site rapid molecular diagnosis.

According to the present invention, a portable isothermal nucleic acid analysis device based on mobile phone detection is provided, including a mobile phone, an excitation optical module, a receiving optical module, a lower isothermal heating unit, an upper constant temperature unit, and a working angle mechanical adjustment unit, wherein:

The mobile phone, in cooperation with the excitation optical module, generates detection light from its built-in flashlight to irradiate and excite the reaction reagents in the reaction test tube. Real-time collection is carried out, and the detection (positive/negative) of the marker to be tested in the sample is realized according to the grayscale information of the collected fluorescence image.

The lower isothermal heating unit is used to provide the temperature environment required for isothermal gene amplification reaction.

The upper layer constant temperature unit is used to cooperate with the lower layer isothermal heating unit to provide a stable temperature gradient between the upper layer and the lower layer required by the reaction reagents in the isothermal gene amplification reaction.

The working angle mechanical adjustment unit is used to produce different reaction effects by adjusting the inclined working angle of the reaction test tube for different detection items or reaction reagents, so as to obtain the best gene amplification reaction efficiency.

In one embodiment, a thermal capacity adjustment unit is also included, wherein:

The heat capacity adjustment unit is located between the lower isothermal heating unit and the upper constant temperature unit, and is kept in close contact with the lower isothermal heating unit and the upper constant temperature unit, and is used to transfer the heat of the lower isothermal heating unit to the upper constant temperature through a custom conduction method unit, and allow it to reach the desired stable temperature

In one embodiment, the heat capacity adjustment unit has a detachable heat dissipation unit, and the heat dissipation unit is composed of equally spaced heat sinks. By adding or reducing the number of heat dissipation units, the heat capacity, heat dissipation area and heat conduction effect can be changed, so that the upper constant temperature unit can achieve the expected working temperature.

In one embodiment, the isothermal heating unit on the lower floor heats the reaction test tube with a cylindrical heating resistance film wrapped by heat-insulating bakelite, and the reaction test tube is inserted into the middle of the cylindrical heating resistance film, and the heating is ensured by the elasticity of the heating resistance film. The tight fit between the reactor and the reaction test tube can obtain excellent heating effect. At the same time, the heater is sealed and wrapped by the heat-insulating Bakelite, which inhibits the heat loss of the heater to the outside and improves the utilization rate of heat.

In one embodiment, it also includes a light source adjustment unit, wherein:

The light source adjustment unit is used to focus the light emitted by the flashlight of the mobile phone onto the reaction test tube.

In one embodiment, a narrow-band filter is arranged between the flashlight of the mobile phone and the light source adjustment unit to filter the original excitation light generated by the flashlight to obtain excitation light within a specific wavelength range.

In one embodiment, a high-pass filter is set between the reaction test tube and the camera of the mobile phone to filter the fluorescent signal received and detected by the mobile phone.

In one embodiment, the working angle mechanical adjustment unit is used to adjust the tilting working angle of the reaction test tube to change the thermal cycle efficiency in the reaction test tube to obtain the optimal nucleic acid isothermal amplification effect according to the reaction requirements of different reaction reagents. Its working angle adjustment resolution is 2°.

In one embodiment, the heating resistance film is provided with a stable working voltage, through which the reaction test tube is heated, and finally the steady-state reaction of the reaction test tube reaches the temperature required for isothermal gene amplification.

In one embodiment, the portable isothermal nucleic acid analysis device based on mobile phone detection can be powered not only by an external power supply, but also by various mobile power sources such as charging treasures, and has the characteristics of portability and strong adaptability to the working environment. .

The invention provides a portable nucleic acid analysis device with simple operation and low cost based on the principle of isothermal amplification by means of mobile phone detection.

Description of drawings

In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the following will briefly introduce the accompanying drawings required in the embodiments or descriptions. Obviously, the accompanying drawings in the following description are only some embodiments of the present invention , for those skilled in the art, other drawings can also be obtained according to these drawings without paying creative labor.

Fig. 1 is the overall structure diagram of a portable isothermal nucleic acid analysis device based on mobile phone detection according to the present invention.

Fig. 2 is a structural diagram of the heat capacity adjustment unit in the present invention.

Fig. 3 is a structural diagram of the middle and lower layer isothermal heating unit of the present invention.

Figure 4.1 is a structural diagram of the working angle mechanical adjustment unit in the present invention.

Fig. 4.2 is another structure diagram of the working angle mechanical adjustment unit in the present invention.

Fig. 5 is a structure diagram of optical detection for fluorescence collection based on mobile phone in the present invention.

Detailed ways

The following will clearly and completely describe the technical solutions in the embodiments of the present invention with reference to the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only some, not all, embodiments of the present invention. The following description of at least one exemplary embodiment is merely illustrative in nature and in no way taken as limiting the invention, its application or uses. Based on the embodiments of the present invention, all other embodiments obtained by persons of ordinary skill in the art without making creative efforts belong to the protection scope of the present invention.

The relative arrangements of components and steps, numerical expressions and numerical values set forth in these embodiments do not limit the scope of the present invention unless specifically stated otherwise.

At the same time, it should be understood that, for the convenience of description, the sizes of the various parts shown in the drawings are not drawn according to the actual proportional relationship.

Techniques, methods and devices known to those of ordinary skill in the relevant art may not be discussed in detail, but where appropriate, such techniques, methods and devices should be considered part of the Authorized Specification.

In all examples shown and discussed herein, any specific values should be construed as illustrative only, and not as limiting. Therefore, other examples of the exemplary embodiment may have different values.

It should be noted that like numbers and letters denote similar items in the following figures, therefore, once an item is defined in one figure, it does not require further discussion in subsequent figures.

Fig. 1 is a schematic diagram of an embodiment of a portable isothermal nucleic acid analysis device based on mobile phone detection in the present invention. As shown in Figure 1, the analysis and diagnosis device includes a mobile phone (1), an excitation optical module (2), a receiving optical module (4), a lower isothermal heating unit (5), an upper constant temperature unit (6), and a working angle mechanical adjustment unit (7).

The lower isothermal heating unit (5) is arranged at the bottom of the reaction reagent (3) and heats the reaction reagent (3), and the upper layer isothermal unit (6) is arranged at the top of the reaction reagent (3) and heats the reaction reagent (3). The capacity adjustment unit (8) is set between the lower isothermal heating unit (5) and the upper constant temperature unit (6), and is kept in close contact with the lower isothermal heating unit (5) and the upper constant temperature unit (6); the working angle is mechanically adjusted The unit (7) includes an optocoupler and a stepping motor at the bottom, the positioning origin is found through the optocoupler, and the lower isothermal heating unit (5), reaction reagent (3), upper constant temperature unit (6) and heat capacity adjustment are driven by the stepping motor Unit (8) moves. The mobile phone (1) is located on the side of the reaction reagent (3) for fluorescence detection.

The flashlight (11), narrow-band filter (101), light source adjustment unit (10), camera (12), and high-pass filter (102) are the unit components of the mobile phone (1), and each component is arranged on the mobile phone (1). ) back.

The excitation optical module (2) includes a light source adjustment unit (10) and a narrow-band filter (101), the narrow-band filter (101) is installed on the light source adjustment unit (10), and the light source adjustment unit (10) is installed on the flashlight (11 )superior;

The receiving optical module (4) includes a high-pass filter (102), and the high-pass filter (102) is installed on the camera (12);

The mobile phone (1) cooperates with the excitation optical module (2), and the flashlight (11) generates detection light to irradiate and excite the reaction reagent (3) in the reaction test tube. At the same time, through the camera (12) and the receiving optical module (4 ) in cooperation with the fluorescent image of the reaction reagent in the reaction test tube is collected in real time, and the detection (positive/negative) of the marker to be tested in the sample is realized according to the grayscale information of the collected fluorescent image.

The lower isothermal heating unit (5) is used to provide the temperature environment required for isothermal gene amplification reaction.

The upper layer constant temperature unit (6) is used to cooperate with the lower layer isothermal heating unit to provide a stable temperature gradient between the upper layer and the lower layer required by the reaction reagents in the isothermal gene amplification reaction.

The working angle mechanical adjustment unit (7) is used for different detection items or reaction reagents, by adjusting the inclined working angle of the reaction test tube to produce different reaction effects, so as to obtain the best gene amplification reaction efficiency.

Based on the portable isothermal nucleic acid analysis device based on mobile phone detection provided by the above-mentioned embodiments of the present invention, the detection light is generated by the flashlight (11) of the mobile phone (1), and the reaction reagent in the reaction reagent (3) is detected by the excitation optical module (2). Irradiation and excitation are performed, and at the same time, the fluorescent image signal transmitted through the receiving optical module (4) is received through the camera (12), and the marker to be detected is detected through the grayscale information. The lower layer isothermal heating unit (5) and the upper layer constant temperature unit (6) cooperate to provide a stable temperature gradient between the upper and lower layers of the reaction reagent; the working angle mechanical adjustment unit (7) is used for different detection items or reaction reagents, through Adjust the inclined working angle of the reaction test tube to produce different reaction effects and obtain the best gene amplification reaction efficiency. Aiming at the isothermal amplification process, the present invention provides a method of using the flash lamp (11) and the camera (12) to replace expensive optical detection devices such as laser, LED, photomultiplier tube, photodiode, etc. The detection method and the principle of isothermal amplification realize a portable accounting and analysis device with simple operation and low cost.

The fluorescence detection device of the present invention also includes a heat capacity adjustment unit (8), wherein:

The heat capacity adjustment unit (8) is used to transfer the heat of the lower layer isothermal heating unit (5) to the upper layer constant temperature unit (6) through a custom conduction method, and make it reach the expected stable temperature; the heat capacity adjustment unit (8) consists of It is composed of a detachable heat dissipation unit (9), and the heat capacity, heat dissipation area and heat conduction effect are changed by adding or reducing the number of heat dissipation units (9), so that the upper constant temperature unit reaches the expected working temperature; the heat dissipation unit (9) is composed of etc. Composed of spaced cooling fins, the heat capacity of the heat capacity adjustment unit (8) can be changed by inserting the number of heat capacity adjustment units (8).

Preferably, the heat dissipation unit has an adjustable surface area, and the heat capacity is changed by adjusting the surface area so that the upper layer constant temperature unit reaches the expected working temperature.

The lower isothermal heating unit (5) heats the reaction test tube by a cylindrical heating resistance film (51) wrapped in heat-insulating bakelite, and the reaction test tube is inserted into the middle of the cylindrical heating resistance film (51). ) elasticity to ensure the tight fit between the heater and the reaction test tube to obtain an excellent heating effect. At the same time, the heater is sealed and wrapped by the thermal insulation bakelite, which inhibits the heat loss of the heater to the outside and improves the use of heat. Rate.

Preferably, the heating resistance film (51) has good elasticity and can be closely attached to the test tube to obtain an excellent heating effect. Wrapping the heater with insulating materials such as bakelite can effectively suppress the external energy loss of the heater and improve The utilization rate of heat, while reducing the interference of the external environment on the heater.

The light source adjustment unit (10) is used to focus the light emitted by the flash lamp (11) onto the reaction reagent (3).

Preferably, the light source emitted by the flash lamp (11) has a relatively large scattering angle, and the light source can be effectively converged by the light source adjustment unit and focused on the reaction test tube.

A narrow-band filter (101) is arranged between the flash lamp (11) and the light source adjustment unit (10), for filtering the original excitation light generated by the flash lamp (11) to obtain excitation light within a specific wavelength range.

Preferably, the excitation center wavelength of the selected fluorescent marker is 492nm, so the narrowband filter (101) allows the light to pass through a wavelength range of 425-500nm, and this wavelength range is used as a detection light source with high energy.

A high-pass filter (102) is arranged between the reaction reagent (3) and the camera (12), for filtering the fluorescent signal received and detected by the mobile phone (1).

Preferably, the central wavelength of the fluorescent signal excited by the selected fluorescent marker is 518nm, so the high-pass filter allows the light to pass through a wavelength range greater than 510nm, which is used to prevent the detection light of the excitation light source from entering the mobile phone (1).

The working angle mechanical adjustment unit (7) is used to meet the reaction requirements of different reaction reagents, by adjusting the inclined working angle of the reaction test tube, changing the thermal cycle efficiency in the reaction test tube, and obtaining the optimal nucleic acid isothermal amplification effect. The adjustment resolution is 2°.

Preferably, the precise positioning of the angle can be realized by a stepping motor and a grating positioning method.

By providing a stable working voltage to the heating resistance film (51), the reaction test tube is heated through it, and finally the steady-state reaction of the reaction test tube reaches the temperature required for isothermal gene amplification.

Preferably, a DC-DC power supply provides a stable working voltage for the heating resistance film (51), and the heating power of the heating resistance film (51) is changed by adjusting the DC-DC output voltage.

This portable isothermal nucleic acid analysis device based on mobile phone detection can be powered not only by an external power supply, but also by various mobile power sources such as charging treasures. It has the characteristics of portability and strong adaptability to the working environment.

A portable isothermal nucleic acid analysis device based on mobile phone detection of the present invention is a portable nucleic acid analysis device with simple operation and low cost based on the principle of isothermal amplification by means of mobile phone detection and natural diffusion temperature gradient formation.

The description of the present invention has been presented for purposes of illustration and description, but is not intended to be exhaustive or limited to the invention in the form disclosed. Many modifications and changes will be apparent to those of ordinary skill in the art. The embodiment was chosen and described in order to better explain the principles of the invention and the practical application, and to enable others of ordinary skill in the art to understand the invention and design various embodiments with various modifications as are suited to the particular use.

Claims (6)

1. the portable nucleic acid analyzer based on mobile phone detection, it is characterised in that：The analytical equipment includes mobile phone (1), excitation Optical module (2) receives optical module (4), lower layer's isothermal heating unit (5), upper layer constant temperature unit (6), operating angle machinery Adjust unit (7)；

Lower layer's isothermal heating unit (5) is arranged in reaction tube (3) bottom and is heated to reaction tube (3), upper layer constant temperature Unit (6) setting at the top of reaction tube (3) and heats reaction tube (3), and thermal capacitance adjusts unit (8) and is arranged in lower layer Between isothermal heating unit (5) and upper layer constant temperature unit (6), and keep and lower layer's isothermal heating unit (5) and upper layer constant temperature list First (6) fit closely；Operating angle mechanical adjustment unit (7) includes the optocoupler and stepper motor of bottom, and it is fixed to be found by optocoupler Position origin drives lower layer's isothermal heating unit (5), reaction tube (3), upper layer constant temperature unit (6) and thermal capacitance by stepper motor Unit (8) is adjusted to move；Mobile phone (1) is located at reaction tube (3) side and carries out fluoroscopic examination to it；The analytical equipment further includes heat Hold and adjust unit (8), thermal capacitance adjusts unit (8) and is used to the heat of lower layer's isothermal heating unit (5) passing through self-defined conduction side Formula is transmitted to upper layer constant temperature unit (6), and reaches expected equilibrium temperature；

Flash lamp (11), spike filter (101), light source adjust unit (10), camera (12), high-pass filter (102) and are The included unit member of mobile phone (1), back of each component setting in mobile phone (1)；

It includes that light source adjusts unit (10) and spike filter (101), spike filter (101) installation to excite optical module (2) It is adjusted on unit (10) in light source, light source adjusts unit (10) and is mounted on flash lamp (11)；

It includes high-pass filter (102) to receive optical module (4), and high-pass filter (102) is mounted on camera (12)；

Mobile phone (1) is matched with excitation optical module (2), is generated detection light by flash lamp (11), is tried being reacted in reaction tube Agent is irradiated, excites, meanwhile, it is matched with optical module (4) is received by camera (12), is tried being reacted in reaction tube The fluorescent image of agent is acquired in real time, and realizes mark to be measured in sample according to the half-tone information of the fluorescent image collected The detection of object；

Lower layer's isothermal heating unit (5) is used to provide the isothermic gene amplification temperature environment that reaction needs；Upper layer constant temperature unit (6) For being matched with lower layer isothermal heating unit, provide in isothermic gene amplification reaction, the upper and lower interlayer needed for reaction reagent is steady Fixed temperature gradient；

Light source adjusts unit (10) and is used to the light that flash lamp (11) is sent out focusing on reaction tube (3)；

The light source angle of scattering that flash lamp (11) is sent out is larger, and can effectively light source be converged by adjusting unit by light source, and be gathered On coke to reaction tube；

A spike filter (101) is set between flash lamp (11) and light source adjust unit (10), for flash lamp (11) the original exciting light generated is filtered, and obtains the exciting light in particular range of wavelengths；

One high-pass filter (102) is set between reaction tube (3) and camera (12), for being received by mobile phone (1) The fluorescence signal of detection is filtered.

2. the portable nucleic acid analyzer according to claim 1 based on mobile phone detection, it is characterised in that：Operating angle Mechanical adjustment unit (7) is used to be directed to different detection project or reaction reagent, by the inclined work angle for adjusting reaction tube It spends to generate different reaction effects, obtains best gene amplification reaction efficiency.

3. the portable nucleic acid analyzer according to claim 1 based on mobile phone detection, it is characterised in that：Thermal capacitance is adjusted For unit (8) by having dismountable heat-sink unit (9) to form, the quantity by adding or reducing heat-sink unit (9) changes thermal capacitance Amount, heat dissipation area and thermal conduction effect so that upper layer constant temperature unit reaches expected operating temperature；The heat-sink unit (9) by Equidistant groups of fins is at the number for adjusting unit (8) by being inserted into thermal capacitance changes the thermal capacitance that thermal capacitance adjusts unit (8).

4. the portable nucleic acid analyzer according to claim 3 based on mobile phone detection, it is characterised in that：Heat-sink unit (9) there is adjustable surface area, change thermal capacitance so that upper layer constant temperature unit reaches expected work by adjusting surface area Temperature.

5. the portable nucleic acid analyzer according to claim 1 based on mobile phone detection, it is characterised in that：Lower layer's isothermal Heating unit (5), the cylindrical shape adding thermal resistance film (51) wrapped up by heat preservation bakelite heat reaction tube, and reaction tube is inserted The centre for entering cylindrical shape adding thermal resistance film (51), by the elasticity of adding thermal resistance film (51) come ensure heater and reaction tube it Between fit closely, obtain excellent heating effect, meanwhile, by keep the temperature bakelite closing package is carried out to heater, it is suppressed that The external heat loss of heater, improves the utilization rate of heat；

Adding thermal resistance film (51) has preferable elasticity, can fit closely to obtain excellent heating effect with test tube, leads to It crosses the thermal insulation materials such as bakelite and the energy dissipation that can effectively inhibit heater external is wrapped up to heater, improve the profit of heat With rate, while reducing interference of the external environment to heater.

6. the portable nucleic acid analyzer according to claim 1 based on mobile phone detection, it is characterised in that：Operating angle Mechanical adjustment unit (7) is used for the reaction requirement for differential responses reagent, by adjusting the inclined work angle of reaction tube, Change the efficiency of cycle in reaction tube, obtain optimal nucleic acid isothermal amplification effect, operating angle adjusts resolution ratio and is 2°；The precise positioning to angle is realized by stepper motor and grating positioning method；

By give adding thermal resistance film (51) provide a stable operating voltage, reaction tube is heated by it, and It is final that the homeostatic reaction of reaction tube is made to reach the temperature needed for isothermic gene amplification；

It is that adding thermal resistance film (51) provides stable operating voltage, and is changed by adjusting DC-DC output voltages by DC-DC power source The heating power of adding thermal resistance film (51).