Microfluidic reagent card and detection method and application thereof

Abstract

A microfluidic reagent card for medical testing and assaying is disclosed. The microfluidic reagent card comprises a card body (10) and a plurality of micropipes, the plurality of micropipes is fixed onto the card body (10). The plurality of micropipes is arranged radially. Each of the micropipe is configured to have a sample inlet end (1) and a closed end (2), and a detection liquid layer (3) and a separation medium layer (4) are arranged inside each micropipe in sequence from the sample inlet end (1) to the closed end (2). The micropipe has a diameter of 0.1-1.0 mm. As the micropipe is configured to have a small diameter, the fluidity of liquid and gel decreases inside the micropipe, and the liquid interface will not flow even when the micropipe is placed in a horizontal direction.

Claims

1. A microfluidic reagent card, wherein comprising a card body and a micropipe, a plurality of the micropipes is fixed on the card body; the micropipe is configured to have a sample inlet end and a closed end, and a detection liquid layer and a separation medium layer are arranged inside the micropipe in sequence from the sample inlet end to the closed end, wherein the separation medium layer is made of gel particles or glass beads, the micropipe has a diameter of 0.1-1.0 mm, wherein a reduced fluidity of a liquid in the micropipe forms a liquid interface holding the separation medium layer in place; the sample inlet end is configured to be connected to an automatic quantitative sample distribution device, and a sample is added into the plurality of the micropipes simultaneously; the automatic quantitative sample distribution device comprises a sample adding component, the sample adding component is connected to a quantitative region, the quantitative region is connected to the sample inlet, a closing portion is provided between the quantitative region and the sample inlet end of the micropipe.

2. The microfluidic reagent card according to claim 1, wherein the detection liquid layer comprises a diluent, the diluent comprises an antibody reagent or does not comprise any antibody reagent.

3. The microfluidic reagent card according to claim 1, wherein the plurality of the micropipes is arranged radially, the closed end of each of the plurality of the micropipes is arranged at the lateral side of the micropipe.

4. The microfluidic reagent card according to claim 1, wherein the automatic quantitative sample distribution device comprises a sample adding component and a plurality of quantitative sample distribution systems.

5. A detection method using the microfluidic reagent card according to claim 1, wherein the detection method comprises the following steps: entering a sample into the micropipe to be mixed with the detection liquid layer in the micropipe, centrifuging the micropipe, and reading a result.

6. A detection method for detecting an antigen-antibody reaction using the microfluidic reagent card according to claim 1, wherein the detection method comprises the following steps: entering a sample into the micropipe to be mixed with the detection liquid layer in the micropipe, centrifuging the micropipe, and reading a result of an antigen-antibody reaction.

7. The detection method for detecting an antigen-antibody reaction using the microfluidic reagent card according to claim 6, wherein the sample and the detection liquid layer are selected for detecting a reaction from a group consisting of blood grouping, antibody screening and cross-matching of blood.

Description

DRAWINGS

(1) FIG. 1 illustrates a micropipe structure of Embodiment 1 of the present invention. Reference number: 1sample inlet end, 2closed end, 3detection liquid layer, 4separation medium layer.

(2) FIG. 2 illustrates a micropipe structure of Embodiment 2 of the present invention. Reference number: 1sample inlet end, 2closed end, 3detection liquid layer, 4separation medium layer, 5 quantitative region, 6 closing portion, 7 exhaust passage.

(3) FIG. 3 illustrates a micropipe structure of Embodiment 3 of the present invention. Reference number: 1sample inlet end, 2closed end, 3micropipe, 5 quantitative region, 6 closing portion, 7exhaust passage, 8sample adding component, 9overflow region.

(4) FIG. 4 illustrates a microfluidic reagent card structure of Embodiment 5. Reference number: 10card body.

(5) FIG. 5 illustrates a microfluidic reagent card structure diagram of Embodiment 6. Reference number: 10card body.

EMBODIMENTS

(6) The following is a detailed description of the preferred embodiments of the present invention, in conjunction with the attached drawings. Experimental methods without specific conditions indicated in the preferred embodiments are usually conducted under normal conditions. The embodiments are provided to better illustrate the specifics of the invention, but they are not limited the invention to the embodiments. Thus, modifications and adaptations to the foregoing embodiments of the present invention conceived by those skilled in the art shall still fall within the scope of the present invention.

(7) Embodiment 1

(8) A micropipe as illustrated in FIG. 1, wherein the micropipe is configured to have a sample inlet end 1 and a closed end 2, a detection liquid layer 3 and a separation medium layer 4 are arranged inside the micropipe in sequence from the sample inlet end 1 to the closed end 2. The micropipe has a diameter of 0.8 mm, and a length of 10 mm.

(9) Separation using the micropipe of the present embodiment: a micropipe gel separation device is used to separate agglutinated from non-agglutinated red blood cells. A predetermined amount of sample is entered into the connected micropipe gel separation device, separating agglutinated from non-agglutinated red blood cells.

(10) When the sample enters into the micropipe and passes through the detection liquid layer, the sample reacts with the detection liquid layer, and then contacts the separation medium layer, which functions to separate and determine the reactants. The small diameter of the micropipe saves the use of sample and reagents, as well as substantially reduces the fluidity of liquid and gel, so that liquid interface does not flow even when the micropipe is placed in a horizontal direction, i.e., the liquid in the capillary can maintain a fixed interface.

(11) Embodiment 2

(12) See Embodiment 1, according to the micropipe shown in FIG. 2, the micropipe is configured to have a sample inlet end 1 and a closed end 2, a detection liquid layer 3 and a separation medium layer 4 are arranged inside the micropipe in sequence from the sample inlet end to the closed end. The micropipe has a diameter of 0.8 mm, and a length of 10 mm. The sample inlet end 1 and a quantitative region 5 are connected. A closing portion 6 is provided between the sample inlet end 1 and the quantitative region 5, and the closing portion 6 is filled with sealing material. After the sample enters into the quantitative region 5, unseal the sealing material. Any material that can function to seal and unseal the closing portion 6 is suitable for the present invention. For example, the present embodiment utilizes paraffin wax to seal the closing portion. After the sample is loaded, the paraffin wax can be melted by warm light, accordingly allowing the sample to enter the micropipe. To ensure the fluidity of the sampling solution, the sample inlet end is configured to have an exhaust passage 7.

(13) The pathway between the quantitative region 5 and the closing portion 6 gradually narrows, the pathway between the closing portion 6 and the sample inlet end 1 is narrower than the pathway in front of the closing portion. The diameter of the pathway in front of the closing portion 6 is 0.08-0.14 mm, the diameter of the pathway between closing portion 6 and sample inlet end 1 is 0.15-0.3 mm. Specifically, the pathway in front of the closing portion 6 has a diameter of 0.1 mm, the pathway between the closing portion 6 and the sample inlet end 1 has a diameter of 0.2 mm.

(14) The micropipe of the present embodiment comprises an automatic quantitative sample distribution device. The micropipe is configured to be connected to the automatic quantitative sample distribution device, which comprises a sample adding component and a plurality of quantitative sample distribution systems. The micropipe gel separation device is used to separate agglutinated from non-agglutinated red blood cells. A predetermined amount of sample enters into the connected micropipe gel separation device, which separates agglutinated from non-agglutinated red blood cells.

(15) Embodiment 3

(16) See Embodiment 2, according to the micropipe shown in FIG. 3, a sample inlet end 1 of the micropipe 3 is connected to an automatic quantitative sample distribution device. The automatic quantitative sample distribution device is connected to a plurality of micropipes 3. The automatic quantitative sample distribution device comprises a sample adding component and a plurality of quantitative sample distribution systems.

(17) Specifically, the present embodiment provides six micropipes 3, which are connected to the automatic quantitative sample distribution device, the sample adding component of the automatic quantitative sample distribution device is configured to take the form of a sample adding region 8, which is connected to the quantitative sample distribution system. The quantitative sample distribution system of the present embodiment comprises six quantitative regions 5 (the number of the quantitative regions correspond to the number of micropipes). The sample adding region 8 is connected to the quantitative region 5. Among the six quantitative regions 5, the furthest from the sample adding region 8 is connected to an overflow region 9. After adding sample into the sample adding region 8, the sample will fill up the six quantitative region 5 from the nearest to the furthest. Once the quantitative region 5 furthest from the sample adding region 8 is filled up, the sample will enter into the overflow region 9. The setting of the overflow region 9 is configured to facilitate easy observation once the overflow region 9 has been filled up once the sample enters into the overflow region 9, it means that the quantitative regions 5 have been fully filled up. Due to the small size of the whole device, in order to avoid any obstruction caused by atmospheric pressure, an exhaust channel 7 is arranged between each two of the quantitative regions 5. Similarly, both the overflow region 9 and the sample inlet end 1 are arranged to have an exhaust channel 7. All of the quantitative regions 5 are configured to have the same size, and to accommodate the same amount of sample, which is to ensure uniformity in the quantities of the experiments. Each quantitative region 5 is connected to the sample inlet end 1 of the micropipe 3 through a pipe. The closing portion 6 is provided in the micropipe, so that the sample and the reagent inside the micropipe do not mix under normal condition.

(18) Embodiment 4 A Human ABO Blood Group Positive Typing and Rh(D) Blood Group Microfluidic Reagent Card

(19) A human ABO blood group positive typing and Rh(D) blood group microfluidic reagent card comprises a card body and a plurality of micropipe, and the plurality of micropipe are fixed onto the card body. The plurality of the micro cubes is divided into 12 groups, each group has 4 micropipes. The plurality of micropipes is arranged radially, and radially distributed according to the center of centrifugation on a plane. The microfluidic reagent card further comprises an automatic quantitative sample distribution device, the micropipe is connected to the automatic quantitative sample distribution device. The automatic quantitative sample distribution device comprises a sample adding component and a plurality of quantitative sample distribution systems. The micropipe gel separation device is used to separate agglutinated from non-agglutinated red blood cells. A predetermined amount of sample enters into the connected micropipe gel separation device, which separates agglutinated from non-agglutinated red blood cells.

(20) The separation medium layer in the micropipe are gel particles.

(21) The detection liquid layers in each group of 4 micropipes are:

(22) TABLE-US-00001 Micropipe No. Component Description Micropipe 1: anti-A reagent anti-A mouse monoclonal (IgM) antibody (clone BRIMA-1) Micropipe 2: anti-B reagent anti-B mouse monoclonal (IgM) antibody (clone LB-2) Micropipe 3: anti-Rh(D) Anti-Rh(D) human monoclonal (IgM) reagent antibody (clone RUM-1) Micropipe 4: antibody diluent Diluent, no antibody
Embodiment 5 Human ABO-positive/Reverse Group and Rh (D) Blood Group Microfluidic Reagent Card

(23) A human ABO-positive/reverse group and Rh (D) blood group microfluidic reagent card, with reference to FIG. 4, comprises a card body and a plurality of micropipes, the plurality of micropipes is fixed on the card body. The plurality of micropipes 12 are divided into 12 groups, each group has 6 micropipes, the plurality of micropipes is arranged radially, and radially distributed according to the center of centrifugation on a plane. The microfluidic reagent card also comprises an automatic quantitative sample distribution device. The micropipe is connected to an automatic quantitative sample distribution device. The automatic quantitative sample distribution device comprises a sample adding component and a plurality of quantitative sample distribution systems. A micropipe gel separation device is used to separate agglutinated from non-agglutinated red blood cells. A predetermined amount of sample enters into the connected micropipe gel separation device, which separates agglutinated from non-agglutinated red blood cells.

(24) The separation medium layer in the micropipe are gel particles.

(25) The detection liquid layers in each group of 6 micropipes are:

(26) TABLE-US-00002 Micropipe No. Component Description Micropipe 1: anti-A reagent anti-A mouse monoclonal (IgM) antibody (clone BRIMA-1) Micropipe 2: anti-B reagent anti-B mouse monoclonal (IgM) antibody (clone LB-2) Micropipe 3: anti-Rh(D) Anti-Rh(D) human monoclonal (IgM) reagent antibody (clone RUM-1) Micropipe 4: control reagent Antibody diluent Micropipe 5, 6: reverse buffer Antibody diluent

(27) Reagent to be used together: as verified, human ABO blood reverse group red blood cell kit (each box contains one A1, B and O reagent, 10 ml/unit) produced by Shanghai blood biomedical Limited Liability Company, is suitable for the microfluidic reagent card of the present invention.

(28) Embodiment 6

(29) FIG. 5 illustrates a microfluidic reagent card, a plurality of micropipes of embodiment 2 are fixed onto a card body 10.

(30) Specifically, there are 6 micropipe groups fixed onto the card body that has a circular shape, each of the 6 micropipe groups has 11 micropipes of embodiment 2. The micropipe groups are arranged in a ring. All of these micropipes are arranged radially, and the closed end of the micropipe is arranged in the lateral side of the micropipe. The micropipes are radially distributed according to the center of centrifugation on a plane.

(31) Embodiment 7 Detection Method of Human ABO Blood Group Positive Typing and Rh(D) Blood Group Microfluidic Reagent Card

(32) A detection method of the human ABO blood group positive typing and Rh(D) blood group microfluidic reagent card according to embodiment 5, comprising the following steps: 1) Take out the microfluidic reagent card, and balance the microfluidic reagent card to room temperature; 2) Mark the microfluidic reagent card, remove the sealing material, avoid cross-contamination, and place the microfluidic reagent card in the centrifugation device; 3) Load sample of whole blood and centrifuge; 4) Read the testing result from a detection window.
Embodiment 8 ABO Blood Group Positive Typing and Rh(D) Blood Group Microfluidic Reagent Card for Newborns

(33) An ABO blood group positive typing and Rh(D) blood group microfluidic reagent card for newborns, with reference to FIG. 4, comprises a card body and a plurality of micropipes, the plurality of micropipes is fixed onto the card body. The plurality of micro cubes is divided into 12 groups, each group has 6 micropipes. The micropipes are arranged radially, and radially distributed according to the center of centrifugation on a plane; the microfluidic reagent card also comprises an automatic quantitative sample distribution device. The micropipe is connected to an automatic quantitative sample distribution device. The automatic quantitative sample distribution device comprises a sample adding component and a plurality of quantitative sample distribution systems. A micropipe gel separation device is used to separate agglutinated from non-agglutinated red blood cells. A predetermined amount of sample enters into the connected micropipe gel separation device, which separates agglutinated from non-agglutinated red blood cells.

(34) The separation medium layer in the micropipe are gel particles;

(35) The detection liquid layers in each group of 6 micropipes are:

(36) TABLE-US-00003 Micropipe No. Component Description Micropipe 1: anti-A reagent anti-A mouse monoclonal (IgM) antibody (clone BRIMA-1) Micropipe 2: anti-B reagent anti-B mouse monoclonal (IgM) antibody (clone LB-2) Micropipe 3: anti-AB reagent anti-B mouse monoclonal (IgM) antibody (clone ES-4) anti-A(B) mouse monoclonal (IgM) antibody (clone ES- 15) Micropipe 4: anti-Rh(D) reagent Anti-Rh(D) human monoclonal (IgM) antibody (clone RUM-1) Micropipe 5: control reagent Antibody diluent Micropipe 6: anti-human Goat anti-human IgG antibody globulin reagent
Embodiment 9 Detection Method of ABO Blood Group Positive Typing and Rh(D) Blood Group Microfluidic Reagent Card for Newborns

(37) As described in Embodiment 8, detection method of ABO blood group positive typing and Rh(D) blood group microfluidic reagent card for newborns comprises the following steps: 1) Take out the microfluidic reagent card, and balance the microfluidic reagent card to room temperature; 2) Mark the microfluidic reagent card, remove the sealing material, avoid cross-contamination, and place the microfluidic reagent card in the centrifugal device; 4) Load sample of whole blood; 5) Centrifuge; 6) Read the testing result from a detection window.
Embodiment 10 A Microfluidic Reagent Card for Human Rh System Typing

(38) A Microfluidic reagent card for human Rh system typing, with reference to FIG. 4, comprises a card body and a plurality of micropipes, the plurality of micropipes is fixed on the card body. The plurality of micro cubes is divided into 12 groups, each group has 6 micropipes. The micropipes are arranged radially, and radially distributed according to the center of centrifugation on a plane. The microfluidic reagent card also comprises automatic quantitative sample distribution device. The micropipe is connected to an automatic quantitative sample distribution device, the automatic quantitative sample distribution device comprises a sample adding component and a plurality of quantitative sample distribution systems, the micropipe gel separation device is used to separate agglutinated from non-agglutinated red blood cells, a predetermined amount of sample enters into the connected micropipe gel separation device, which separates agglutinated from non-agglutinated red blood cells.

(39) The separation medium layer in the micropipe are gel particles.

(40) The detection liquid layers in each group of 6 micropipes are:

(41) TABLE-US-00004 Micropipe No. Component Description Micropipe 1: anti-C reagent Anti-C antibody human monoclonal (IgM) antibody (clone MS-24) Micropipe 2: anti-c reagent Anti-c antibody human monoclonal (IgM) antibody (clone MS-33) Micropipe 3: anti-D reagent Anti-Rh(D) human monoclonal (IgM) antibody (clone RUM-1) Micropipe 4: anti-E reagent Anti-E antibody human monoclonal (IgM) antibody (clone MS-80/MS-258) Micropipe 5: anti-e reagent Anti-e antibody human monoclonal (IgM) antibody (clone MS-16/MS-21/MS-63) Micropipe 6: control reagent Antibody diluent
Embodiment 11 Detection Method of a Microfluidic Reagent Card for Human Rh System Typing

(42) A detection method of a microfluidic reagent card for human Rh system typing according to embodiment 10 comprises the following steps: 1) Take out the microfluidic reagent card, and balance the microfluidic reagent card to room temperature; 2) Mark the microfluidic reagent card, remove the sealing material, avoid cross-contamination, and place the microfluidic reagent card in the centrifugal device; 4) Load sample of whole blood; 5) Centrifuge; 6) Read the testing result from a detection window.

(43) Finally, please note that, the embodiments above are intended to illustrate the specifics of the invention only, with no intention to limit the scope of the invention to these embodiments. Although the invention has been described in detail with references to the preferred embodiments above, a person skilled in the art would know that modifications or equivalent replacements without departing from the spirit and scope of the present invention shall still fall within the claimed scope of the present invention.