Patent Application
20230123106
The present invention relates generally to a bioassay device, and more particularly, to provide a bioassay device including all units used in bioassay and a bioassay system with the bioassay device with fully automated complex bioassay operation process.
Enzyme-Linked Immunosorbent Assay (ELISA) or Enzyme-linked immunoassay (EIA) is a specific antigen-antibody reaction test. The specific binding characteristics of the smear can detect the sample waiting to be tested, and in combination with the enzyme to carry out a color reaction, which can show the existence of a specific antigen or antibody, and can use the depth of the color for quantitative analysis, so as to achieve the purpose of detection and screening.
A biochip is a miniature device that utilizes biomaterials that can produce specific biochemical reactions with the biomolecules to be tested on a substrate, and can be quantified by a highly sensitive detection system. The biochip provides the advantages of low-cost bioanalytical testing capabilities. In molecular biology, biochips are basically miniaturized substrates that can perform hundreds or thousands of biochemical reactions simultaneously.
However, conventional ELISA is to use large and expensive equipment to receive optical or electronic signals to detect the status of biochemical molecular reactions after performing complex bioassay procedures, such as observation with a microscope and additional photography device to capture the screen for further analysis, so it takes a certain amount of time and manual operation. On the other hand, conventional biochips need to be additionally equipped with other expensive and large-scale image capture systems or equipment to detect and capture the luminescent images of the biochips after the bioassay process for subsequent analysis.
With the increasing popularity of the concept of point of care testing (POCT), that is, a personalized health test with a short analysis time and simple operation, in order to overcome the traditional biomolecular detection method, which requires large-scale equipment and complex bioassay process, it is imperative to develop more sensitive, simpler testing equipment and methods.
A primary objective of the present invention is to provide a bioassay device on which all units that will be used in bioassay are included.
Another objective of the present invention is to provide a bioassay system, which fully automates the complex bioassay operation process.
In order to achieve the aforementioned objectives, the present invention provides a bioassay device, which includes a main body, a biomolecular image sensor, and a first electrical connection portion. The main body defines a sensing groove. The biomolecular image sensor is disposed in the sensing groove and includes an image sensing unit. The first electrical connection portion is disposed at one side of the main body and is electrically connected to the biomolecular image sensor.
In a preferred embodiment, the diameter of the sensing groove tapers from the top to the bottom, and the biomolecular image sensor is disposed at the bottom of the sensing groove.
In a preferred embodiment, the bioassay device further includes a magnetic unit disposed in the main body and below the sensing groove.
In a preferred embodiment, the main body defines a reaction groove for accommodating a reactant.
In a preferred embodiment, the main body defines at least one reagent groove for accommodating a reagent.
In a preferred embodiment, the main body defines at least one first accommodating groove for accommodating a pipette tip.
In a preferred embodiment, the main body defines at least one second accommodating groove for accommodating a water absorbing unit.
In order to achieve the aforementioned objectives, the present invention provides a bioassay device, which includes a main body, a biomolecular image sensor, and a first electrical connection portion. The main body includes a cover body and a box body and defines a sensing groove, a reaction groove and at least one reagent groove, the cover body is disposed on the box body, and the sensing groove, the reaction groove and the at least one reagent groove all penetrate the top of the cover body and the top of the box body, the reaction groove is used for accommodating a reactant, and at least one reagent groove is used for accommodating a reagent. The biomolecular image sensor is disposed in the sensing groove and includes an image sensing unit. The first electrical connection portion is disposed at one side of the box body, and is electrically connected to the biomolecular image sensor.
In order to achieve the aforementioned objectives, the present invention provides a bioassay system, including a casing, a control device, a mounting seat, a robotic arm, a pipette nozzle, and a bioassay device. The control device is disposed inside the casing. The mounting seat is disposed inside the casing, and has a second electrical connection portion. The second electrical connection portion is electrically connected to the control device. The robotic arm is disposed inside the casing and is electrically connected to the control device. The pipette nozzle is disposed on the robotic arm and is electrically connected to the control device. The bioassay device includes a main body, a biomolecular image sensor and a first electrical connection portion. The main body defines a sensing groove. The biomolecular image sensor is disposed in the sensing groove and includes an image sensing unit. The first electrical connection portion is disposed at one side of the main body, and is electrically connected to the biomolecular image sensor and the second electrical connection portion.
In a preferred embodiment, the bioassay system further includes a first magnet, and the first magnet is disposed on the mounting seat.
The effect of the present invention is that the bioassay device of the present invention can dispose all the units that will be used in bioassay on the main body, so as to form a special set for bioassay.
Furthermore, the bioassay system of the present invention can fully automate the complex bioassay operation process, so as to achieve the following advantages: first, it reduces manual operation steps, avoids human interference, and can shorten the detection time; second, reduce the volume and weight of the system and reduce costs; third, improve the sensitivity to achieve the detection effect of a single molecule; fourth, expand the scope of application; fifth, avoid the waste of specimens and minimize the amount of specimens used; sixth, achieve the goal of using the same small amount of sample to perform ultra-trace detection of multiple target molecules in a single detection.
The present invention will be apparent to those skilled in the art by reading the following detailed description of a preferred embodiment thereof, with reference to the attached drawings, in which:
The accompanying drawings are included to provide a further understanding of the invention, and are incorporated in and constitute a part of this specification. The drawings illustrate embodiments of the invention and, together with the description, serve to explain the principles of the invention.
As shown in
As shown in
As shown in
Specifically, the box body 120 has a first side 120, a second side 1202, a third side 1203, and a fourth side 1204. The top part of the first side 1201 protrudes to form a protruding platform 1201A, a concave portion 1201B is formed inward below the protruding platform 1201A, and the surface of the concave portion 1201B is further recessed to form a bonding groove 1201C. The sensing groove 11 is close to the center of the main body 10, and the diameter of the sensing groove 11 tapers from the top to the bottom; more specifically, the sensing groove 11 forms an inverted tapered portion 111 in the box body 120, and an opening 112 is defined at the bottom of the inverted tapered portion 111. The reaction groove 12 and the reagent grooves 131-133 penetrate through the top of the protruding platform 1201A. The first accommodating grooves 141-148 are respectively located on both sides of the sensing groove 11 and are disposed in two rows, wherein one row of the first accommodating grooves 141-144 is close to the second side 1202 of the box body 120, and the other row of the first accommodating grooves 145-148 are located close to the fourth side 1204 of the box body 120. The second accommodating grooves 16 are respectively located between the sensing groove 11 and the reagent grooves 131-133 and between the sensing groove 11 and the reaction groove 12. The second side 1202 and the fourth side 1204 of the box body 120 respectively have a plurality of anti-slip bars 18 and a plurality of positioning blocks 19.
Preferably, the length of the cover body 110 is 100 mm, the width of the cover body 110 is 60 mm, the length of the box body 120 is 68 mm, the width of the box body 120 is 60 mm, and the total of the height of the cover body 110 and the height of the box body 120 is 93 mm.
Preferably, the first water-absorbing units 171 and the second water-absorbing units 172 can be made of absorbent cotton, cloth, paper, sponge or water-absorbing resin and other materials with water-absorbing capacity. However, it is not limited to thereto, and any material with water absorbing ability can be the first water absorbing units 171 and the second water absorbing units 172 of the present invention.
As shown in
As shown in
Specifically, the carrier 25 can be a microparticle, and through the EDC/NHS reaction, the biomolecule 24 to be tested, such as an antibody, is linked with the microparticle by forming an amide bond; the antibody, the microparticle and the sample to be tested are then mixed, so as to grab the antigen to be tested from the sample to be tested; then the secondary antibody of the modified biotin is combined with the antigen; the biotin is combined with streptavidin to bring a plurality of horseradish peroxidase (HRP) molecule to form a complex of microparticle-antibody-antigen-antibody-biotin-streptavidin-polyHRP. The complex is the reactant of the first biomolecular image sensor 20A.
Preferably, the microparticles are preferably magnetic beads of 1-3 μm, and the magnetic beads may use magnetic elements, such as iron (Fe), nickel (Ni), cobalt (Co), etc., neodymium iron boron (Nb—Fe—B) and other ferromagnetic alloys, or magnetic materials of Fe3O4, Fe2O3, FeO. Alternatively, non-magnetic materials such as Au, sepharose, polystyrene, and SiO2 can also be used for the microparticles.
Preferably, only a single carrier 25 needs to be accommodated in a single microstructure 221 as much as possible, so the size and height of the microstructures 221 need to match the size and height of the carrier 25. More specifically, the diameter of the microstructure 221 is preferably 1.3-1.8 times larger than the diameter of the microparticles, the depth is preferably 1.2-1.3 times larger than the diameter of the microparticles, and the aspect ratio is preferably 1-1.2 times. With 2 μm beads, the micropores are about 2.5-3 μm in diameter and about 2.5 μm deep. The distance between the micropores is about 2-3 pm. For example, if the carrier 25 is a microparticle with a diameter of 2 μm, and the microstructures 21 can be grooves with a diameter of 2.5-3 μm and a depth of 2.5 μm, and the spacing is 2-3 μm.
As shown in
Preferably, the image sensing unit can be a CMOS image sensor manufactured by using a semiconductor process, and after the packaging is completed, its surface is oxidized to form a light-penetrating flat surface composed of SiO2. The flat surface is the protective layer at the light-receiving surface for receiving incident light. Next, a plurality of detection molecules 21 is directly fixed on the light-receiving surface by means of chemical modification, for example, by using oxygen atoms in SiO2 on the light-receiving surface, firstly with 3-aminopropyl, APTES silane compounds for surface-modifying to have amine groups (NH2) on the surface, so as to interact with detection molecules 21 such as antibodies, receptor proteins, DNA, aptamers, or other chemical molecules to create bonds to directly immobilize the detection molecules 21 on the light-receiving surface of the image sensing unit. More specifically, the EDC/NHS reaction can be used to make the carboxyl group on the antibody as the detection molecule 21 bond with the amine group on the light-receiving surface, so as to be immobilized on the light-receiving surface; or, glutaraldehyde can be used to connect protein G and the amine group on the light-receiving surface. Since protein G can bind to the Fc region of most antibodies, it can flexibly change the specific antibody as detection molecule 21 according to different biomolecules to be tested. The antibody is mixed with the sample to be tested to capture the antigen to be tested from the sample to be tested; then the secondary antibody of the modified biotin is combined with the antigen; the biotin is combined with streptavidin to bring in multiple horseradish peroxidase (HRP) molecules; to form antibody-antigen-antibody-biotin-streptavidin-polyHRP complex. The complex is the reactant of the second biomolecular image sensor 20B.
In a preferred embodiment, the biomolecular image sensor 20 is used for biological or chemical analysis, such as detecting the presence and/or concentration of a biomolecule 24 in a sample to be tested; that is, the incident light can be a light emitted by the fluorescent label, reporter molecule label, or chemiluminescent label of the biomolecule to be tested, and more specifically, the biomolecule 24 to be tested by the biomolecular image sensor 20, in the process of a biological or chemical analysis, can react with other molecules and emit incident light such as chemiluminescence or fluorescence. Moreover, the biomolecules 24 to be tested can be, for example, proteins, peptides, antibodies, nucleic acids, etc. According to the present invention, the fluorescent label can be, but not limited to, FITC, HEX, FAM, TAMRA, Cy3, Cy5, quantum dot, etc., and can be used with a quencher dye.
When the biomolecule 24 to be tested binds to the chemiluminescent label, the reactant at least includes the biomolecule 24 to be tested in the sample to be tested, the reaction groove 12 accommodates the reactant, and the three reagent grooves 131-133 accommodate phosphate buffered saline (PBS), Luminol, also known as photosensitizer, and peroxide respectively.
When the biomolecule 24 to be tested binds to the fluorescent label, the reactant at least includes the biomolecule 24 to be tested in the sample to be tested, the reaction groove 12 accommodates the reactant, the reagent groove 131 accommodates reagents such as phosphate buffered saline; the remaining reagent grooves 132-133 are not injected with any reagents and can even be omitted.
In a preferred embodiment, the image sensing unit 21 may be a backside illuminated Complementary Metal-Oxide-Semiconductor (CMOS) image sensor or a front illuminated CMOS image sensor, but the invention is not limited thereto.
In a preferred embodiment, the unit pixel 211 includes a photoelectric conversion unit (not shown), and the photoelectric conversion unit may be a unit that generates and accumulates electrons corresponding to incident light. For example, the unit pixel 211 may be a photodiode, a photo transistor, a photo gate, a pinned photo diode (PPD), an avalanche photodiode (APD), single-photon avalanche diode (SPAD), photomultiplier tube (PMT), or any combination thereof.
In some embodiments, the biomolecular image sensor 20 includes a glass substrate (not shown) and the image sensing unit 21, and the image sensing unit 21 is disposed on the glass substrate.
As shown in
As shown in
Before starting the bioassay, first, as shown in
As shown in
In step S100, the reactants are uniformly mixed in phosphate buffered saline. More specifically, the control device 2 controls the movement of the robotic arm 4, and the robotic arm 4 controls the movement of the pipette nozzle 5. Step S100 further includes: the pipette nozzle 5 takes out the pipette tips 151-154 from the first accommodating grooves 141-144; the pipette nozzle 5 uses the pipette tips 151-154 to suck phosphate buffered saline from the reagent groove 131; the pipette nozzle 5 uses the pipette tips 151-154 to add phosphate buffered saline into the reaction groove 12 to form a reaction solution; the first magnet 9 moves to the bottom of the protruding platform 1201A, and the first magnet 9 provides a magnetic force to adsorb the reactant onto the side wall of the reaction groove 12, and the pipette tips 151-154 are utilized by the pipette nozzle 5 to draw phosphate buffered saline from the reaction groove 12; the pipette tips 151-154 are placed by the pipette nozzle 5 in the first accommodating grooves 141-144; the first magnet 9 is removed, so that the reactants fall off from the side walls of the reaction groove 12.
In step S101, the reactant and phosphate buffered saline are mixed into a reaction solution and injected into the sensing groove 11. Step S101 further includes: the pipette nozzle 5 takes out the pipette tip 155 from the first accommodating groove 145 and uses the pipette tip 155 to suck phosphate buffered saline from the reagent groove 131; the phosphate buffered saline is injected into the reaction groove 12, and the reactants are suspended in phosphate buffered saline to form a reaction solution; the pipette nozzle 5 uses the pipette tip 155 to suck the reaction solution and inject the reaction solution into the sensing groove 11, and the reactants are easy to follow the sidewall of the inverted tapered portion 111 to flow onto the surface of the biomolecular image sensor 20.
In step S102, the reactants fall into the microstructures 221. More specifically, the means for the carrier 25 to fall into the microstructures 221 include the following two: (1) the control device 2 controls the vibration device 8 to generate vibration, and the vibration provided by the vibration device 8 is transmitted to the bioassay device 6 through the mounting seat 3, and the carrier 25 is affected by the vibration and falls into the microstructures 221; (2) the second magnet 42 is located directly under the sensing groove 11, the second magnet 42 provides a magnetic force, and the carrier 25 is subjected to the magnetic force and falls into the microstructures 221 under the influence. In other words, if the bioassay system is equipped with the vibration device 8, the bioassay device 6 may not be equipped with the magnetic unit 40; if the bioassay system is not equipped with the vibration device 8, the bioassay device must be equipped with the magnetic unit 40; only one of the two options is needed so as to save costs.
Step S103, removing the phosphate buffered saline in the sensing groove 11. Step S103 further includes: the pipette nozzle 5 uses the pipette tip 155 to suck phosphate buffered saline from the sensing groove 11;
Step S104, preparing a chemiluminescent solution (ECL solution). Step S104 further includes: the pipette nozzle 5 takes out the pipette tip 156 from the first accommodating groove 146; the pipette nozzle 5 uses the pipette tip 156 to suck luminol from the reagent groove 132; luminol is injected into reagent groove 133 and forms a chemiluminescent solution with peroxide. It is worth noting that the pipette nozzle 5 can also be moved to the reagent groove 133 to absorb the peroxide, and then the peroxide is injected into the reagent groove 132 to form a chemiluminescent solution with luminol.
In step S105, the chemiluminescent solution is injected into the sensing groove 11. Step S105 further includes: the pipette nozzle 5 uses the pipette tip 156 to suck the chemiluminescent solution; the pipette nozzle 5 uses the pipette tip 156 to inject the chemiluminescent solution into the sensing groove 11, and the chemiluminescent solution easily follows the sidewall of the inverted tapered portion 111 to flow onto the surface of the biomolecular image sensor 20. The chemiluminescent solution can make the chemiluminescent label of the biomolecule 24 to be tested emit light.
In step S106, the chemiluminescent solution in the sensing groove 11 is removed. Step S106 further includes: the pipette nozzle 5 draws the chemiluminescent solution from the sensing groove 11 by using the pipette tip 156;
In step S107, each of the unit pixels 211 respectively detects an incident light in a single microstructure 221, the incident light including the light emitted by the chemiluminescent label of the biomolecule 24 to be tested.
In step S108, the incident light received by each of the unit pixels 211 is transmitted through the unit pixels 211 to generate an electron.
In step S109, a voltage signal is generated according to the electrons through the readout circuit 23, and the voltage signal is transmitted to the control device 2 through the first electrical connection portion 30 and the second electrical connection portion 301.
In step S110, the control device 2 analyzes whether the biomolecule 24 to be tested exists or not according to the voltage signal.
In step S111, if the biomolecule 24 to be tested exists, the control device 2 further compares it with the standard concentration curve to obtain the concentration of the biomolecule 24 to be tested.
In step S112, the control device 2 analyzes whether the signal reading value exceeds the threshold value according to the voltage signal.
In step S113, the control device 2 defines the unit pixel whose measured signal reading value exceeds the threshold value as 1.
In step S114, the control device 2 defines the unit pixel whose measured signal reading value does not exceed the threshold value as 0.
In step S115, the control device 2 calculates the total number of unit pixels as 1 and compares it with the standard concentration curve to obtain the concentration of the biomolecule 24 to be tested.
In some embodiments, the bioassay system can be not configured with the vibration device 8 and the first magnet 9, the bioassay device may not be configured with the magnetic unit 40, and the reagent groove contains a surfactant. Step S102 in these embodiments includes: the pipette nozzle 5 takes out the pipette tip 157 from the first accommodating groove 147; the pipette nozzle 5 uses the pipette tip 157 to suck the surfactant from the reagent groove; the surfactant is injected into the sensing groove 11, and the surfactant easily flows along the sidewall of the inverted tapered portion 111 onto the surface of the biomolecular image sensor 20; the pipette nozzle 5 places the pipette tip 157 into the first accommodating groove 147. Thereby, the surfactant can significantly reduce the surface tension of the reaction solution, so that the carrier 25 can fall into the microstructures 221 smoothly. Compared with the vibration force or the magnetic force, reducing the surface tension of the reaction solution can increase the ratio of the carrier 25 falling into the microstructures 221. Furthermore, the bioassay system omits the vibrating device 8 and the first magnet 9 and the bioassay device 6 omits the magnetic unit 40, so that the manufacturing cost can be further reduced, and the volume and weight can also be reduced.
It is worth noting that, during the movement of the pipette nozzle 5, the pipette tips 151-158 can first contact the second water absorbing units 172 in the second accommodating grooves 16, and the second water absorbing units 172 in the second accommodating grooves 16 can absorb the liquid attached to the outer walls or tips of the pipette tips 151-158, so as to prevent the liquid from dripping into the reagent grooves 131-133 and contaminating the reagents. After the pipette tips 151-158 return to the first accommodating grooves 141-148, the first water absorbing units 171 in the first accommodating grooves 141-148 can absorb the pipette tips 151-158 to prevent the residual liquid from volatilizing into the casing 1 and contaminating the reagent.
As shown in
The present invention utilizes chemiluminescence or photoluminescence for the second biomolecular image sensor 20B to detect biomolecules. The difference from the foregoing method is that steps S100 and S102 are omitted.
In summary, the bioassay device of the present invention can set all the units that will be used in bioassay on the main body, so as to form a bioassay-specific set.
Furthermore, the bioassay system of the present invention can fully automate the complex bioassay operation process, so as to achieve the following advantages: first, it reduces manual operation steps, avoids human interference, and can shorten the detection time; second, reduce the volume and weight of the system and reduce costs; third, improve the sensitivity to achieve the detection effect of a single molecule; fourth, expand the scope of application; fifth, avoid the waste of specimens and minimize the amount of specimens used; sixth, achieve the goal of using the same small amount of sample to perform ultra-trace detection of multiple target molecules in a single detection.
Although the present invention has been described with reference to the preferred embodiments thereof, it is apparent to those skilled in the art that a variety of modifications and changes may be made without departing from the scope of the present invention which is intended to be defined by the appended claims.
This application claims priority of U.S. provisional application No. 63/255,446, filed on Oct. 14, 2021, the content of which is incorporated herein in its entirety by reference.
| Number | Date | Country | |
|---|---|---|---|
| 63255446 | Oct 2021 | US |
