TWI398636B - Detecting method of liquid sample - Google Patents

Description

Fluid detection method

The present invention relates to a method for quantitatively detecting a fluid, and more particularly to a method for quantitatively detecting a fluid used in biochemical detection and immunoassay.

In the conventional technique of performing biochemical detection and immunodetection using a fluid detecting test piece, the fluid detecting test piece is designed with a flow path or a micro flow path structure on the substrate or the substrate, and the surrounding of the flow path is not a water absorbing material, and is to be tested. The fluid is mostly composed of a highly viscous composition such as protein or saccharide. Therefore, when the fluid to be tested flows, it will remain on the flow path, so that the fluid to be tested cannot be completely reacted, thus not only causing the fluid to be tested. The waste is more likely to cause errors in the final test results.

In addition, the fluid detecting test piece of the prior art can be designed with a micro-flow channel structure in fluid transfer, and utilizes the capillary phenomenon generated by the micro-flow path structure to passively transfer the fluid through the flow path to the reaction detecting area; The method is to apply a driving force to the fluid when the fluid to be tested is injected, that is, using a pressure or the like, so that the fluid can actively pass through the flow path to reach the reaction detecting area. However, in either of the above manners, after the fluid to be tested is injected into the flow channel, bubbles of different sizes are often generated to block the flow path, causing an error in actual measurement, and even causing the test to fail.

Finally, in the test strips of the prior art, the flow path or the micro-channel structure is formed on the substrate by using die casting or injection molding, so polyethylene (PE) and polyvinyl chloride (PVC) must be used. Or a higher-priced plastic polymer such as polypropylene (PP) is used as the material, which in turn causes an increase in the overall cost of the test piece.

In order to overcome the above disadvantages, the present invention provides a fluid detecting method, which mainly comprises the following steps:

(1) Providing a substrate with at least a first-class track recessed from the upper surface thereof. The flow channel comprises a first fluid zone, a second fluid zone and a third fluid zone connected in sequence, a nitrocellulose layer is formed at the bottom of the second fluid zone and the third fluid zone, and the nitrocellulose layer comprises a hollow mesh configuration Wherein the second fluid zone is for fluid transfer and the third fluid zone is for fluid reaction, therefore, the average thickness of the nitrocellulose layer of the second fluid zone is no greater than the thickness of the third fluid zone nitrocellulose layer, and the third fluid zone The nitrocellulose layer absorbs a quantity of fluid. In addition, a reactive material is formed in the hollow network configuration of the nitrocellulose layer;

(2) injecting a fluid from the first fluid region of the substrate to allow fluid to enter the third fluid region via the second fluid region;

(3) causing the nitrocellulose layer of the third fluid zone to absorb a quantity of fluid;

(4) Detecting by forming an optical reaction by interaction of a specific component in the fluid with a reaction material of the third fluid zone.

The main object of the present invention is to provide a fluid detecting method in which a flow path of a substrate is provided with a water absorbing nitrocellulose layer, and since a water volume per unit volume of nitrocellulose is constant, it can be set on a substrate. The volume of the nitrocellulose layer provides quantitative detection of the fluid.

Another object of the present invention is to provide a fluid detecting method in which a flow path of a substrate having a hollow network configuration has a nitrocellulose layer, and bubbles in the fluid flow through the hollow network configuration. It will be destroyed, so that bubbles can be prevented from blocking the flow path, providing stable and reliable detection results.

Since the present invention discloses a method for quantitatively detecting a fluid, the physical, chemical, and solution coating techniques utilized therein are well known to those of ordinary skill in the art, and therefore, the description below will not be fully described. At the same time, the drawings in the following texts are indicative of the features related to the features of the present invention, and are not required to be completely drawn according to the actual situation.

Referring to FIG. 1, a preferred embodiment of the present invention is a flow of a quantitative method for detecting a fluid, which is used for detecting a specific component in a fluid, and mainly comprises the following steps:

Step 1: First, a substrate 10 is provided. Referring to FIG. 2, the substrate 10 is recessed from the upper surface 100 thereof to at least the first channel 11, and the flow channel 11 includes the first fluid region 111 and the second fluid region sequentially connected. 112 and a third fluid zone 113. In a preferred embodiment, substrate 10 is a biocompatible material. Please continue to refer to Figure 3, which is a cross-sectional view along line AA of Figure 2. A nitrocellulose layer 1121 and 1131 having a hollow network configuration is formed at the bottom of the second fluid region 112 and the third fluid region 113, wherein the second fluid region 112 is for fluid transfer, and the third fluid region 113 is for fluid supply. The reaction. The average thickness Da of the nitrocellulose layer 1121 of the second fluid zone is not greater than the thickness Db of the nitrocellulose layer 1131 of the third fluid zone, and the amount of absorbed liquid of the nitrocellulose layer 1131 of the third fluid zone is fixed. Further, the hollow mesh configuration of the nitrocellulose layers 1121 and 1131 includes a reaction material, and the composition of the reaction material is related to the type of the component to be tested contained in the fluid.

Step 2: Injecting a fluid L (not shown) from the first fluid zone 111 of the substrate 10 to cause the fluid L to reach the third fluid zone 113 after being injected into the first fluid zone 111 via the second fluid zone 112.

Step 3: The nitrocellulose layer 1131 of the third fluid zone 113 is caused to absorb a quantity of the fluid L.

Step 4: detecting a reaction signal by interacting with a specific component of the fluid L and the reaction material of the third fluid region 113, wherein the reaction signal may be a cold light reaction signal, a fluorescent reaction signal, or a light absorption. Reaction signal, or electrochemical reaction signal.

Further, in order to reduce the influence of the capillary action between the flow path and the fluid, the flow path proposed by the present invention is not a so-called micro flow path of the prior art, and the width Wa of the second fluid region 112 and the third fluid region 113 It is preferably at least 0.3 mm with Wb.

In the production, the manner in which the nitrocellulose layers 1121 and 1131 are formed and the manner in which the reaction material is formed are as follows. First, a nitrocellulose powder is mixed with an organic solvent containing an ester and a ketone to form a nitrocellulose solution; and the nitrocellulose solution is cast in the second fluid region 112 and the first The bottom of the three-fluid zone 113, after drying, forms a nitrocellulose layer 1121 at the bottom of the second fluid zone 112, and forms a nitrocellulose layer 1131 at the bottom of the third fluid zone 113. For better casting, the surface roughness (Ra value) of the flow path 11 is preferably between 3 and 50 microns.

After the nitrocellulose solution is dried, a nitrocellulose layer having a hollow network configuration is formed. In order to adjust a preferred hollow network configuration, the nitrocellulose powder of the nitrocellulose solution of the present invention is mixed with an organic solvent containing an ester and a ketone. The preferred volume ratio is 1:9. Since the water absorption per unit volume of the nitrocellulose is constant, the volume of the corresponding nitrocellulose solution can be derived from the volume of the fluid to be absorbed, and then cast. This makes it possible to fix the volume of the liquid required for detection and to be suitable for micro-testing.

After the nitrocellulose layers 1121 and 1131 are respectively dried and formed at the bottoms of the second fluid region 112 and the third fluid region 113, the reaction solution containing the reaction material is injected into the nitrocellulose layers 1121 and 1131, and subjected to air drying or lyophilization. Thereafter, it remains in the form of a powder in the nitrocellulose layers 1121 and 1131.

The manner in which the nitrocellulose layer 1121, 1131 and the reaction material are formed is formed by sequentially forming a nitrocellulose layer and then injecting the reaction material, and the reaction solution containing the reaction material may be added to the nitrocellulose powder. (nitrocellulose powder) in a nitrocellulose solution consisting of an organic solvent containing an ester and a ketone; after mixing, the mixed solution is cast in the second fluid zone 112 and the third The bottom of the fluid zone 113 is subjected to an air drying or freeze drying process, and the nitrocellulose solution is formed into the nitrocellulose layers 1121 and 1131, and the reaction material is powdered and left in the nitrocellulose layers 1121 and 1131.

Due to the different components to be tested, the reactions required for the detection are also different; and depending on the type of reaction, various signals are generated. For example, in the biochemical test, the enzyme is used to catalyze the test substance and the chemical reagent in the fluid, thereby generating a specific signal for detection. Therefore, for biochemical testing, the reaction material will contain enzymes and corresponding chemical reagents. On the other hand, if certain proteins in the sample, such as the presence of a fetal protein (-fetoprotein), are detected, the specific antibody is used to specifically bind to the protein to be tested, and then other chemical reagents are used. The antibody that has been bound to the protein to be tested is reacted to emit a signal for detection. Therefore, an immunoassay is required, and the reaction material contains immunological reagents such as chemicals and antibodies. Therefore, the substrate 10 provided by the present invention can be used for detecting various components to be tested in various biological samples (such as urine, blood, and the like).

The preferred embodiment described above uses a substrate having three fluid regions, and according to the method for quantitatively detecting a fluid according to the present invention, the substrate used may further be provided with a fourth fluid region after the third fluid region of the flow channel. (not shown) for storing excess fluid in the flow path. The configuration and formation mode of the nitrocellulose layer in the fourth fluid zone, the composition and preferred ratio of the nitrocellulose solution used, and the composition of the reaction material are the same as those of the preferred embodiment described above, and the detailed description thereof will not be repeated here.

The above description is only for the preferred embodiment of the present invention, and is not intended to limit the present invention. The above description is to be understood by those skilled in the art, and thus the other embodiments are not disclosed. Equivalent changes or modifications made in the spirit of the invention are to be included in the scope of the claims below.

1, 2, 3, 4. . . step

10. . . Substrate

100. . . Upper surface

11. . . Runner

111. . . First fluid zone

112. . . Second fluid zone

113. . . Third fluid zone

1121, 1131. . . Nitrocellulose layer

Da. . . Nitrifying fiber layer 1121 thickness

Db‧‧‧Nitrocellulose layer 1131 thickness

Wa‧‧‧Width of the second fluid zone

Wb‧‧‧ width of the third fluid zone

1 is a flow chart of a fluid detecting method in accordance with a preferred embodiment of the present invention.

2 is a schematic view of a substrate provided by a fluid detecting method according to a preferred embodiment of the present invention.

3 is a cross-sectional view of a substrate provided by a fluid detecting method according to a preferred embodiment of the present invention.

1, 2, 3, 4. . . step

Claims (36)

A method for quantitatively detecting a fluid for detecting a specific component in a fluid, comprising: providing a substrate, the substrate recessing at least a first-class channel from an upper surface thereof, the flow channel comprising a first fluid region sequentially connected, a second fluid zone and a third fluid zone, the second fluid zone and the bottom of the third fluid zone are formed with a nitrocellulose layer, the nitrocellulose layer comprising a hollow network configuration, wherein the second fluid zone is for fluid Transmitting, the third fluid zone is for reacting with a fluid, the average thickness of the nitrocellulose layer of the second fluid zone is not greater than the thickness of the nitrocellulose layer of the third fluid zone, and the nitrocellulose layer of the third fluid zone is capable of absorbing quantitative a fluid, a reactive material is formed in the hollow network configuration of the nitrocellulose layer; a fluid is injected from the first fluid region of the substrate, and the fluid enters the third fluid region via the second fluid region; The nitrocellulose layer of the three fluid zone absorbs a quantity of fluid; and is detected by a reaction of a specific component of the fluid with a reactive material of the third fluid zone to form a reaction signal. The method for quantitatively detecting a fluid according to claim 1, wherein the average thickness of the nitrocellulose layer of the second fluid zone is smaller than the thickness of the nitrocellulose layer of the third fluid zone. The method for quantitatively detecting a fluid according to claim 2, wherein the nitrocellulose layer is formed by casting a nitrocellulose solution after casting to the bottom of the second fluid zone and the third fluid zone and drying. A method for quantitatively detecting a fluid according to the third aspect of the invention, wherein the nitrocellulose solution is formed by mixing an organic solvent containing a ketone and an ester with a nitrocellulose powder. A method for quantitatively measuring a fluid according to claim 4, wherein a preferred volume ratio of the nitrocellulose powder to an organic solvent containing a ketone and an ester is 1:9. A method for quantitatively measuring a fluid according to claim 2, wherein the second fluid zone and the third fluid zone have a minimum width of 0.3 mm. A method for quantitatively detecting a fluid according to claim 2, wherein the substrate is a biocompatible material. A method for quantitatively measuring a fluid according to claim 2, wherein the flow path has a surface roughness Ra of between 3 micrometers and 50 micrometers. The method for quantitatively detecting a fluid according to claim 3, wherein the reaction material is injected into the nitrocellulose layer by a reaction solution, and then formed into a powder after drying. A method for quantitatively measuring a fluid according to claim 9 wherein the drying process is freeze drying. A method for quantitatively measuring a fluid according to claim 9 wherein the drying process is air drying. The method for quantitatively detecting a fluid according to claim 3, wherein the reaction material is injected into the nitrocellulose solution by a reaction solution, and the nitrocellulose solution is simultaneously formed into a nitrocellulose layer by a drying process, and the reaction material is formed into a powder form. . A method for quantitatively measuring a fluid according to claim 12, wherein the drying process is freeze drying. A method for quantitatively measuring a fluid according to claim 12, wherein the drying process is air drying. A method for quantitatively detecting a fluid according to claim 2, wherein the reaction material is a chemical and an enzyme reagent. A method for quantitatively measuring a fluid according to claim 2, wherein the reaction material is an antibody and a chemical reagent. The method for quantitatively measuring a fluid according to claim 1, wherein the average thickness of the nitrocellulose layer of the second fluid zone is equal to the thickness of the nitrocellulose layer of the third fluid zone. The method for quantitatively measuring a fluid according to claim 17, wherein the flow path further comprises a fourth fluid zone, and a bottom of the fourth fluid zone is also formed with a nitrocellulose layer, the nitrocellulose layer comprising a hollow mesh structure For the storage of excess fluid. The method for quantitatively measuring a fluid according to claim 18, wherein the nitrocellulose layer is cast with a nitrocellulose solution after being cast in the second fluid zone, the third fluid zone and the fourth fluid zone, and then dried. Formed. A method for quantitatively measuring a fluid according to claim 19, wherein the nitrocellulose solution is The nitrocellulose powder is formed by mixing an organic solvent containing an ester and a ketone. A method for quantitatively measuring a fluid according to claim 20, wherein a preferred volume ratio of the nitrocellulose powder to an organic solvent containing an ester and a ketone is 1:9. The method of quantitatively detecting a fluid according to claim 18, wherein the second fluid zone and the third fluid zone have a minimum width of 0.3 mm. A method for quantitatively measuring a fluid according to claim 18, wherein the substrate is a biocompatible material. A method for quantitatively measuring a fluid according to claim 18, wherein the flow path has a surface roughness Ra of between 3 micrometers and 50 micrometers. The method for quantitatively detecting a fluid according to claim 19, wherein the reaction material is injected into the nitrocellulose layer by a reaction solution, and then formed into a powder after drying. A method for quantitatively measuring a fluid according to claim 25, wherein the drying process is freeze drying. A method for quantitatively measuring a fluid according to claim 25, wherein the drying process is air drying. The method for quantitatively detecting a fluid according to claim 19, wherein the reaction material is injected into the nitrocellulose solution by a reaction solution, and the nitrocellulose solution is simultaneously formed into a nitrocellulose layer by a drying process, and the reaction material is formed into a powder. . A method for quantitatively measuring a fluid according to claim 28, wherein the drying process is freeze drying. A method for quantitatively measuring a fluid according to claim 28, wherein the drying process is air drying. A method for quantitatively measuring a fluid according to claim 18, wherein the reaction material is a chemical and an enzyme reagent. The method for quantitatively detecting a fluid according to claim 18, wherein the reaction material is an antibody and a chemical reagent. The method for quantitatively detecting a fluid according to Item 2 or Item 17 of the patent application, wherein the reaction signal is a cold light reaction signal. A method for quantitatively detecting a fluid according to claim 2 or 17, wherein the reaction signal is a fluorescent reaction. The method for quantitatively detecting a fluid according to Item 2 or Item 17 of the patent application, wherein the reaction signal is a light absorption reaction signal. The method for quantitatively detecting a fluid according to Item 2 or Item 17 of the patent application, wherein the reaction signal is an electrochemical reaction signal.