Chemiluminescence characteristic detection method

Abstract

The invention discloses a chemiluminescence characteristic detection method. The chemiluminescence characteristic detection method comprises the following steps: 1, adding a sample; and 2, mixing and reacting the enzyme label, the magnetic beads and the sample. And 3, transferring the sample liquid containing the magnetic beads into a detection tube. Sample liquid in the detection tube is sucked in a mode of adsorbing the magnetic beads by a magnetic field, and the magnetic beads are retained in the detection tube. And 4, cleaning the magnetic beads in the detection tube for one or more times. And 5, carrying out light-emitting detection. According to the method, the positions of the magnetic beads are fixed through magnetic attraction on the outer side of the detection tube, and the cleaning liquid is attracted away under the condition that the magnetic beads are not affected, so that the magnetic beads are reserved in the same detection tube in multi-step cleaning, and compared with a cleaning method that the magnetic beads are sucked into an attraction head and transferred in the prior art, the cleaning efficiency is improved. According to the invention, the magnetic bead loss caused by magnetic bead transfer residue is obviously reduced.

Description

Technical Field

[0001] The present invention belongs to the technical field of chemiluminescence characteristic detection, and in particular relates to a chemiluminescence characteristic detection method. Background Art

[0002] In the detection of chemical characteristics of a single person, it is necessary to separate the magnetic beads from the sample liquid and wash the magnetic beads multiple times. The patent with publication number CN110514653A provides a chemiluminescence analyzer and its use method, which sucks the magnetic beads and liquid in the hole into the TIP tip together during the magnetic bead washing process. When the magnetic beads in the transfer tip need to be separated, the magnetic suction motor is turned on to push the magnetic suction plate close to the transfer tip. The magnetic suction needle groove matches the transfer tip. The magnetic suction plate has an adsorption effect on the magnetic beads, so that the magnetic beads are retained in the transfer tip, and the liquid in the transfer tip flows out to achieve the separation of the magnetic beads. Then the magnetic beads in the TIP tip are transferred to the next hole for washing or subsequent testing. However, this separation method requires the magnetic beads to be sucked into the TIP tip and transferred multiple times; there may be residual magnetic beads in each suction and transfer process; and then after multiple washings, a large loss of magnetic beads is caused, which reduces the accuracy of the detection. Summary of the invention

[0003] The purpose of the present invention is to provide a chemiluminescence characteristic detection method.

[0004] The present invention provides a chemiluminescence characteristic detection method, comprising the following steps: Step 1: Add the sample to the sample well of the reagent strip.

[0005] Step 2: Mix the enzyme label and magnetic beads on the reagent strip with the sample and react for a preset time.

[0006] Step 3: Transfer the sample liquid containing magnetic beads obtained in step 2 to the test tube. Then, apply a magnetic field to the inner cavity of the test tube so that the magnetic beads in the test tube are adsorbed on the inner wall of the test tube; absorb the sample liquid in the test tube, and the magnetic beads remain in the test tube.

[0007] Step 4: Wash the magnetic beads in the test tube once or multiple times. During each washing process:

[0008] Step 4-1: Inject the cleaning solution into the detection tube containing magnetic beads and mix well.

[0009] Step 4-2: Apply a magnetic field to the inner cavity of the detection tube so that the magnetic beads in the detection tube are adsorbed on the inner wall of the detection tube. The liquid in the detection tube is sucked out, and the magnetic beads remain in the detection tube.

[0010] Step 5: Add substrate to the detection tube, use a photomultiplier tube to detect the luminescence of the detection tube, and obtain the chemiluminescence characteristic detection result of the sample.

[0011] Preferably, the magnetic field is applied to the inner cavity of the detection tube by placing a permanent magnet close to one side of the outer wall of the detection tube so that the magnetic beads are simultaneously adsorbed on one side of the inner cavity of the detection tube.

[0012] Preferably, the side of the permanent magnet facing the detection tube is provided with an arc-shaped groove matching the shape of the detection tube; when a magnetic field is applied to the inner cavity of the detection tube, the arc-shaped groove on the permanent magnet fits with the cylindrical outer side of the detection tube.

[0013] Preferably, in step 3 and step 4-2, the pipette tip is moved to a side of the central axis of the detection tube away from the permanent magnet before aspirating the liquid.

[0014] Preferably, a flat bottom area is provided at the bottom of the inner cavity of the detection tube.

[0015] Preferably, the magnetic field is applied to the inner cavity of the detection tube by using two permanent magnets or two different positions of a permanent magnet close to the two sides of the outer wall of the detection tube, so that the magnetic beads are adsorbed on both sides of the inner cavity of the detection tube at the same time.

[0016] Preferably, the magnetic field is applied to the inner cavity of the detection tube by using a ring magnet to be placed from bottom to top on the outside of the detection tube, so that a magnetic field with magnetic flux lines along the radial direction of the detection tube is generated in the detection tube, so that the magnetic beads are evenly adsorbed on the inner wall of the detection tube.

[0017] Preferably, the magnetic field is applied to the inner cavity of the detection tube by energizing an electromagnet close to the outer side of the detection tube.

[0018] Preferably, in step one, the reagent strip to which the sample is added is heated and incubated.

[0019] Preferably, in step 4-1, the cleaning solution containing magnetic beads in the detection tube is repeatedly sucked and released multiple times by using a pipette tip, so that the magnetic beads and the cleaning solution are mixed evenly.

[0020] Preferably, after step 3 and step 4-2 are executed, the magnetic field applied to the inner cavity of the detection tube is removed.

[0021] The present invention has the following beneficial effects: 1. The present invention fixes the position of the magnetic beads by magnetic attraction on the outside of the detection tube, and absorbs the cleaning solution without affecting the magnetic beads, so that the magnetic beads are retained in the same detection tube during multi-step cleaning. Compared with the cleaning method of sucking the magnetic beads into the suction head and transferring them in the prior art, the present invention significantly reduces the loss of magnetic beads caused by the residual magnetic beads during transfer.

[0022] 2. The magnetic bead separation process of the present invention only requires two steps: magnetic suction and sample liquid aspiration. Compared with the prior art operations of aspirating the magnetic bead mixture, magnetically aspirating the tip, releasing the liquid from the tip, and transferring the magnetic beads in the tip to the next well, the operation process is simplified. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 It is a side schematic diagram of the relative position of the permanent magnet and the detection tube in the present invention.

[0024] Figure 2 It is a top view schematic diagram of the relative positions of the permanent magnet and the detection tube in the present invention.

[0025] Figure 3 It is a schematic diagram of the process of separating magnetic beads and liquid in a detection tube in the present invention. DETAILED DESCRIPTION

[0026] The present invention will be further described below in conjunction with the accompanying drawings.

[0027] Example 1 like Figure 1 , Figure 2 and Figure 3 As shown, a magnetic bead separation and cleaning method for chemiluminescence characteristic detection comprises the following steps: Step 1: Magnetic bead separation stage.

[0028] 1-1. Use a pipette tip to transfer the reacted sample solution containing magnetic beads to the detection tube.

[0029] 1-2. Attach a permanent magnet to the outer side of the detection tube and apply a magnetic field to the inner cavity of the detection tube, so that the magnetic beads in the detection tube are adsorbed on the inner wall of the detection tube.

[0030] In some embodiments, an arc-shaped groove matching the shape of the detection tube is provided on the side of the permanent magnet facing the detection tube, so that the detection area of ​​the permanent magnet and the detection tube is increased, so that magnetic beads at different positions can be stably adsorbed on the inner wall of the detection tube.

[0031] 1-3. Use the pipette tip to absorb the sample liquid in the test tube. The magnetic beads in the test tube remain in the test tube under the action of magnetic force.

[0032] In some optional embodiments, the tip of the pipette is located at the side of the bottom of the suction detection tube away from the permanent magnet, so as to increase the distance between the pipette and the magnetic beads, further prevent the magnetic beads from being sucked into the pipette, and reduce the loss of magnetic beads. In a further optional embodiment, a flat bottom area with a diameter of 2 mm is provided at the bottom of the inner cavity of the detection tube, so as to ensure that the liquid in the detection tube can be fully sucked out when the pipette deviates from the center of the detection tube.

[0033] 1-4. The pipette tip transfers the sample liquid to the waste liquid storage area outside the test tube. At the same time, the permanent magnet is moved away from the test tube so that the magnetic beads in the test tube can flow freely.

[0034] Step 2: Magnetic bead cleaning stage: Repeat the use of cleaning solution to clean the magnetic beads in the detection tube multiple times; the cleaning process is the same each time.

[0035] The process of performing a cleaning is as follows: 2-1. Use a pipette tip to absorb the unused cleaning solution and inject it into the test tube.

[0036] 2-2. Use the pipette tip to repeatedly suck up and release the cleaning solution containing magnetic beads in the detection tube to fully mix the magnetic beads and the cleaning solution so that the cleaning solution can fully clean the magnetic beads.

[0037] 2-3. Reattach the permanent magnet to the outer side of the detection tube and apply a magnetic field to the inner cavity of the detection tube so that the magnetic beads in the detection tube are adsorbed on the inner wall of the detection tube.

[0038] 2-4. The pipette tip transfers the absorbed cleaning liquid to the waste liquid storage area outside the detection tube. At the same time, the permanent magnet is kept away from the detection tube so that the magnetic beads in the detection tube can flow freely.

[0039] After multiple washes, the sample liquid remaining on the inner wall of the test tube and the magnetic beads is effectively removed, and since the magnetic beads remain in the test tube, there is almost no loss.

[0040] Example 2 A magnetic bead separation and cleaning method for chemiluminescence feature detection, this embodiment differs from embodiment 1 only in the way of applying a magnetic field to the inner cavity of a detection tube; in this embodiment, no movable permanent magnet is provided, but an electromagnet is attached to the outer surface of the detection tube, and in steps 1-2 and 2-3, the generation or disappearance of the magnetic field in the inner cavity of the detection tube is controlled by turning the electromagnet on and off.

[0041] Example 3 A magnetic bead separation and cleaning method for chemiluminescence characteristic detection, this embodiment is different from embodiment 1 only in the way of applying a magnetic field to the inner cavity of the detection tube; in this embodiment, two permanent magnets are used to move to the left and right sides of the detection tube respectively, so that the magnetic beads are adsorbed to the two sides of the inner cavity of the detection tube. The suction head is inserted from the center of the detection tube and sucks the waste liquid without magnetic beads.

[0042] In this embodiment, the adsorption area of ​​the magnetic beads is increased to ensure that all the magnetic beads can be effectively and firmly adsorbed.

[0043] Example 4 A magnetic bead separation and cleaning method for chemiluminescence characteristic detection, this embodiment is different from embodiment 1 only in the way of applying a magnetic field to the inner cavity of the detection tube; in this embodiment, a ring magnet is used to be placed on the detection tube from bottom to top, so that a magnetic field with magnetic flux lines along the radial direction is generated in the detection tube, and the magnetic beads are uniformly adsorbed on the inner wall of the detection tube. The suction head is inserted from the center of the detection tube and sucks the waste liquid without magnetic beads.

[0044] Compared with Example 3, the adsorption area of ​​the magnetic beads is further increased in this embodiment to ensure that all the magnetic beads can be effectively and firmly adsorbed.

[0045] Example 5 A chemiluminescence characteristic detection method comprises the following steps: Step (1): Add the sample to the sample well of the reagent strip.

[0046] In this embodiment, the reagent strip is provided with a sample well, two lysis wells, a substrate well, three wash wells, an enzyme label well, a magnetic bead well, and a detection tube arranged in sequence. In addition to the sample well, in some other embodiments, the number of lysis wells and wash wells can be increased or decreased (including completely eliminated) according to actual needs.

[0047] Step (2): Install the TIP tip on the pipette and heat the reagent strip to improve the detection reaction efficiency.

[0048] Step (3): Puncture the aluminum film sealing all the wells of the reagent strip. Use a TIP pipette tip to transfer the reagents in different wells of the reagent strip to perform sample reaction.

[0049] The following is a detailed description of the sample reaction processing process: The sample well, two lysis wells, one substrate well, three wash wells, one enzyme label well, one magnetic bead well, and one detection tube are marked as wells 1 to 10, as shown in the following table: 1 2 3 4 5 6 7 8 9 10 Sample well Cracking hole Cracking hole Substrate well Washing fluid hole Washing fluid hole Washing fluid hole ELISA well Magnetic Bead Hole Detection hole The reaction process without lysis is: adding enzyme label, adding magnetic beads and aspirating and mixing for 4 minutes, transferring the sample mixture to the detection tube, separating and cleaning the magnetic beads, and adding substrate. The process of separating and cleaning the magnetic beads is the same as that in Example 1, and the number of cleanings is three times. The reagent transfer process is: (1→8), (9→8), (8→10)*, (5→10)*, (6→10)*, (7→10)*, (4→10).

[0050] The reaction process under the condition of one lysis is: one lysis treatment and suction mixing, adding enzyme label, adding magnetic beads and suction mixing reaction for 4 minutes, transferring the sample mixture to the detection tube, magnetic bead separation and cleaning, and adding substrate. The process of magnetic bead separation and cleaning is the same as that in Example 1, and the number of cleanings is three times. The reagent transfer process is: (1→3), (8→3), (9→3), (3→10)*, (5→10)*, (6→10)*, (7→10)*, (4→10).

[0051] The reaction process under the condition of secondary lysis is: two lysis treatments and suction mixing, adding enzyme label, adding magnetic beads and suction mixing reaction for 4 minutes, transferring the sample mixture to the detection tube, magnetic bead separation and cleaning, and adding substrate. The process of magnetic bead separation and cleaning is the same as that in Example 1, and the number of cleanings is three times. The reagent transfer process is: (1→3), (2→3), (8→3), (9→3), (3→10)*, (5→10)*, (6→10)*, (7→10)*, (4→10).

[0052] In the above process, * indicates that after the reagent transfer is completed, the liquid in the detection hole needs to be sucked out using a tip under magnetic suction conditions; (5→10)*, (6→10)*, (7→10)* correspond to three washing operations.

[0053] Step (4): Use the detection port of the photomultiplier tube to press against the top opening of the detection tube, perform luminescence detection on the substrate mixed with the sample magnetic beads in the detection tube, and obtain the chemiluminescence characteristic detection result of the sample.

Claims

1. A chemiluminescence characteristic detection method, characterized in that: The following steps are involved: Step 1: Add the sample to the sample well of the reagent strip; Step 2: Mix the enzyme label and magnetic beads on the reagent strip with the sample and react for a preset time; Step 3: Transfer the sample liquid containing magnetic beads obtained in step 2 to a detection tube; then, apply a magnetic field to the inner cavity of the detection tube so that the magnetic beads in the detection tube are adsorbed on the inner wall of the detection tube; aspirate the sample liquid in the detection tube, and the magnetic beads remain in the detection tube; Step 4: Wash the magnetic beads in the test tube once or multiple times; during each washing process: Step 4-1: inject the cleaning solution into the detection tube containing magnetic beads and mix well; Step 4-2, applying a magnetic field to the inner cavity of the detection tube so that the magnetic beads in the detection tube are adsorbed on the inner wall of the detection tube; aspirating the liquid in the detection tube, and the magnetic beads remain in the detection tube; Step 5: Add substrate to the detection tube, use a photomultiplier tube to detect the luminescence of the detection tube, and obtain the chemiluminescence characteristic detection result of the sample.

2. A chemiluminescence characteristic detection method according to claim 1, characterized in that: The magnetic field is applied to the inner cavity of the detection tube by using a permanent magnet close to one side of the outer wall of the detection tube.

3. A chemiluminescence characteristic detection method according to claim 2, characterized in that: The side of the permanent magnet facing the detection tube is provided with an arc groove matching the shape of the detection tube; when a magnetic field is applied to the inner cavity of the detection tube, the arc groove on the permanent magnet fits with the cylindrical outer side of the detection tube.

4. A chemiluminescence characteristic detection method according to claim 2, characterized in that: In step 3 and step 4-2, the pipette tip is moved to the side of the central axis of the detection tube away from the permanent magnet before aspirating the liquid.

5. A chemiluminescence characteristic detection method according to claim 4, characterized in that: The inner cavity bottom of the detection tube is provided with a flat bottom area.

6. A chemiluminescence characteristic detection method according to claim 1, characterized in that: The magnetic field is applied to the inner cavity of the detection tube by using two permanent magnets or two different positions of one permanent magnet close to the two sides of the outer wall of the detection tube respectively.

7. A chemiluminescence characteristic detection method according to claim 1, characterized in that: The magnetic field is applied to the inner cavity of the detection tube by using a ring magnet to be placed from bottom to top on the outside of the detection tube, so that a magnetic field with magnetic flux lines along the radial direction of the detection tube is generated in the detection tube.

8. A chemiluminescence characteristic detection method according to claim 1, characterized in that: The magnetic field is applied to the inner cavity of the detection tube by energizing an electromagnet close to the outer side of the detection tube.

9. A chemiluminescence characteristic detection method according to claim 1, characterized in that: In step 1, the reagent strip with the sample added is heated and incubated.

10. A chemiluminescence characteristic detection method according to claim 1, characterized in that: In step 4-1, the cleaning solution containing magnetic beads in the detection tube is repeatedly sucked and released by using a pipette tip to mix the magnetic beads and the cleaning solution. After step 3 and step 4-2 are executed, the magnetic field applied to the inner cavity of the detection tube is removed.

Images