Chemiluminescence ELISA kit for detecting concentration of drug tacrolimus in human whole blood and Chemiluminescence bottom indicator

A chemiluminescence enzyme and chemiluminescence technology, which is applied in the field of medical detection, can solve the problem that the luminescence performance cannot meet the sensitivity and stability of the detection requirements, affect the sensitivity and stability of the detection kit, and affect the monitoring results of the patient's blood drug concentration, etc. problem, to achieve the effect of shortening the detection time, high accuracy and long duration

Active Publication Date: 2013-07-31
同昕生物技术(北京)有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] Chemiluminescent chromogenic substrates have an important impact on the effect of chemiluminescent immunoassays, and directly affect the sensitivity and stability of the detection kit. A good chemiluminescent chromogenic substrate can make the detection limit of the detection system reach the picogram level, or even higher. Chemicals with high and general effects not only have low sensitivity and cannot meet the test requirements, but also have extremely poor stability, which makes the results of each test vary greatly, which directly affects the monitoring results of the patient's blood drug concentration, causing doctors to be unable to use the test results according to each test. Make the correct judgment on the patient's medication
At present, the luminescence display substrate liquid commercially available in China is used in plate-type chemiluminescence detection kits, and its luminescence performance often fails to meet the sensitivity and stability required for detection.

Method used

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  • Chemiluminescence ELISA kit for detecting concentration of drug tacrolimus in human whole blood and Chemiluminescence bottom indicator
  • Chemiluminescence ELISA kit for detecting concentration of drug tacrolimus in human whole blood and Chemiluminescence bottom indicator
  • Chemiluminescence ELISA kit for detecting concentration of drug tacrolimus in human whole blood and Chemiluminescence bottom indicator

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1. Preparation and use of a kit in which tacrolimus monoclonal antibody is the coating source and the marker is an enzyme-labeled hapten

[0027]1. The detection principle of the kit using tacrolimus monoclonal antibody as the coating source and the enzyme-labeled tacrolimus hapten as the marker is as follows:

[0028] The microwell strip is pre-coated with tacrolimus monoclonal antibody, and the tacrolimus in the sample and the enzyme-labeled tacrolimus compete with the pre-coated anti-tacrolimus antibody on the microwell strip , so that the absorbance value of the sample is negatively correlated with the content of tacrolimus contained in the sample, and the amount of tacrolimus in the sample can be obtained by comparing with the standard curve.

[0029] 2. Kit composition:

[0030] Kit No. I:

[0031] 1. Chemiluminescent microtiter plates coated with a coating source: the coating source is a mouse monoclonal antibody against tacrolimus, and the coating conc...

Embodiment 2

[0054] Weigh 1 mg of the above hapten and dissolve it in 2 ml of N, N-dimethylformamide, stir at room temperature for 30 min, add dropwise to 10 ml (5 mg) of horseradish peroxidase solution, stir overnight at room temperature, and use Sephadex G -25 for chromatographic purification. Example 2. Application of a kit using tacrolimus monoclonal antibody as a coating source and an enzyme-labeled hapten as a marker

[0055] The kit of the invention is used for quantitatively detecting the drug concentration of tacrolimus in human whole blood.

[0056] 1. Pretreatment of human whole blood samples

[0057] Shake the human whole blood tacrolimus standard and quality control solution and samples to be tested stored in the refrigerator at 2-8°C, fully shake and mix on the shaker, and immediately pipette 150 μl of each sample into the corresponding centrifuge tube accurately, After adding an equal volume of sample pretreatment solution, cap the tube immediately, rotate at high speed wi...

Embodiment 3

[0091] Example 3. The original coating is a conjugate of tacrolimus and carrier protein, and the enzyme-labeled secondary antibody is an enzyme-labeled kit

[0092] The working principle of this kit is:

[0093] When the coating on the microwell strip of the chemiluminescent microtiter plate is originally a conjugated tacrolimus hapten and carrier protein, after adding the standard solution or sample solution to the microwell of the chemiluminescent microtiter plate, add the antibody The specific antibody of tacrolimus, the tacrolimus in the sample competes with the tacrolimus-coupled antigen on the chemiluminescence microtiter plate, and then the enzyme-labeled anti-antibody is added for amplification, and the substrate is used for color development In this way, the absorbance value of the sample is negatively correlated with the content of tacrolimus, and the content of tacrolimus in the sample can be obtained according to the standard curve obtained from the standard.

[0...

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Abstract

The invention provides a chemiluminescence ELISA kit for detecting the concentration of the drug tacrolimus in human whole blood, which belongs to the field of medical detection reagents. The kit comprises a chemiluminescence enzyme label plate and enzyme markers, wherein, coating antigens on the chemiluminescence enzyme label plate are monoclonal antibodies against tacrolimus, secondary antibodies or semiantigens of tacrolimus coupled with carrier proteins; the invention is characterized in that: the kit further comprises self-made chemiluminescence substrate chromogenic reagent which enables the kit to have a higher signal to noise ratio and detection sensitivity, and the enzyme markers are horseradish peroxidase marked antibodies, antigens or antiantibodies. The kit is applicable to an open type semi-automatic chemiluminescence measuring instrument and to an automatic measuring system, accords with current situation in China, costs little, has a detection effect hardly different from that of an imported reagent, enables mass and rapid detection to be realized and is easier to popularize and apply.

Description

technical field [0001] The invention relates to medical detection technology, in particular to a detection kit for tacrolimus drug concentration in human whole blood. Background technique [0002] Tacrolimus is a macrolide immunosuppressant discovered by Fujisawa Company in 1984. It was first used clinically in 1989 and has been widely used in the treatment of rejection after transplantation. Its mechanism of action is that after tacrolimus binds to its binding protein FKBP12, it inhibits the activity of phosphatase, thereby inhibiting nuclear regulatory proteins and T cell activating factors (such as interleukin-2), proto-oncogenes and corresponding receptors of cytokines ( Such as the expression of interleukin-2 receptor). However, the actual situation is that the individual absorption and clearance rates of tacrolimus vary greatly, and there is no close correlation between the dosage and the blood concentration of tacrolimus. There is a certain degree of toxicity at high...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/76G01N33/577G01N33/535
Inventor 贾凤芹焦守恕李全
Owner 同昕生物技术(北京)有限公司
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