Application of kit for detecting D-dimer by spatial proximity chemiluminescence method

A detection kit, chemiluminescence technology, used in chemiluminescence/bioluminescence, analysis by chemical reaction of materials, measurement devices, etc., can solve the problem of unstable detection results, poor repeatability, and many influencing factors, etc. problems, to achieve the effect of eliminating luminous interference, easy to fully automate, and simple production process

Pending Publication Date: 2021-10-19
无锡壹闪生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Its operation is complicated and there are many influencing factors, resulting in unstable test results and poor repeatability

Method used

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Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0029] 1. Preparation of calibrator

[0030] (1) Preparation of calibration product dilution: Weigh 14.1 g of potassium dihydrogen phosphate, sodium dihydrogen phosphate (2H 2 O) 3.0g, add ultrapure water to dissolve, Proclin-3000.5 ~ 1mL, after mixing, add ultrapure water to make up to 1000mL, obtain the calibration product dilution, store at 2 ~ 8 ℃ for later use.

[0031] (2) Preparation of calibrator: the concentration of the calibrator is 0, 10ng / mL, 50ng / mL, 500ng / mL, 1000ng / mL, 10000ng / mL, and the purified D-dimer is diluted to the corresponding concentration with the calibrator diluent , Store at 2-8°C for later use.

[0032] 2. Preparation of enzyme markers

[0033] (1) Weigh 5 mg of HRP and dissolve it in 1 mL of distilled water, add 0.2 mL of freshly prepared 0.1 M sodium periodate solution to the supernatant, stir at room temperature in the dark for 20 min, put the above solution into a dialysis bag, and test for 1 mM pH4. 4 sodium acetate buffer dialyzed overni...

Embodiment 1

[0051] This embodiment provides a D-dimer detection kit, including: enzyme markers, luminescent markers, auxiliary agents, triggers and calibrator; wherein, the calibrator includes no less than 6 different concentrations of D-Dimer Antigen calibrator and 0.1M phosphate buffer; enzyme markers include peroxidase-labeled D-Dimer detection antibody and 0.05M phosphate buffer; luminescent markers include 9,10-dihydroacridine-labeled D - Dimer capture antibody and 0.05M Tris buffer; the components of the auxiliary agent include luminescent auxiliary agent and pH 6.0 citrate buffer; the trigger agent is 0.05M pH 8.0 Tris-HCl buffer.

[0052] 1. Preparation of calibrator

[0053] (1) Preparation of calibration product dilution: Weigh 14.1 g of potassium dihydrogen phosphate, sodium dihydrogen phosphate (2H 2 O) 3.0g, add ultrapure water to dissolve, Proclin-3000.8mL, after mixing, add ultrapure water to make up to 1000mL to obtain the calibration product dilution, store at 4°C for la...

Embodiment 2

[0068] This embodiment provides a method for detecting D-dimer using a D-dimer detection kit in spatially adjacent chemiluminescence, including steps:

[0069] S1: Add 25 μL of standard, 25 μL of peroxidase-labeled D-Dimer detection antibody and 25 μL of luminescent marker to the chemiluminescence reaction tube.

[0070] S2: React in a 37°C incubator for 15 minutes.

[0071] S3: Add 5 μL of auxiliary agent to the chemiluminescent reaction tube, shake and mix, and let it stand for 1-2 minutes; then add 75 μL of trigger agent, shake and mix, detect immediately, and read the signal value.

[0072] S4: Carry out logistic four-parameter fitting on the concentration and luminescence value of the calibrator, and calculate the sample concentration through the luminescence value of the sample.

[0073] The present invention adopts spatial proximity chemiluminescence method to detect D-dimer, and the sensitivity can reach 10 ng / mL; D-dimer has a diagnostic specificity of 86.3% for acut...

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Abstract

The invention relates to application of a kit for detecting D-dimer by a spatial proximity chemiluminescence method. The kit comprises an enzyme marker, a luminous marker, an auxiliary agent, a triggering agent and a calibrator, wherein the calibrator comprises calibrators of D-Dimer antigens with different concentrations and a 0.1 M phosphate diluent; the enzyme marker comprises the following components: a peroxidase-labeled D-Dimer detection antibody and a 0.05 M phosphate buffer solution; the luminous marker is prepared from the following components: a D-Dimer capture antibody marked by 9, 10-dihydroacridine and a 0.05 M Tris buffer solution; the auxiliary agent comprises a luminous auxiliary agent and a citrate buffer solution; and the 0.05 M Tris buffer solution is selected as a triggering agent. As a real homogeneous chemiluminescence technology, the detection method provided by the invention does not need a carrier, does not have coating and washing processes, has few components in the kit, and is high in sensitivity, good in repeatability and simple to operate.

Description

[0001] This application is a divisional application with an application date of September 25, 2018, an application number of 201811115058.5, and an invention title of "a kit for the detection of D-dimer by spatially adjacent chemiluminescence and its detection method". technical field [0002] The invention relates to the field of biotechnology, in particular to the application of a kit for detecting D-dimers by space proximity chemiluminescence method. Background technique [0003] D-dimer (D-Dimer) is a degradation product of cross-linked fibrin, and its formation mechanism is: under the action of thrombin, fibrinogen removes polypeptide A and polypeptide B from the α chain and β chain in turn, forming Fibrin I and fibrin II, fibrin is cross-linked on the blood vessel wall under the action of factor XII, and is cleaved by activated plasmin to produce various FDP fragments. Due to the cross-linking of the γ-chain, two D fragments connected by the γ-chain are produced, that ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/76G01N33/577G01N33/535
CPCG01N21/76G01N33/535G01N33/577
Inventor 奚伟红朱丹丹廖鸳鸯
Owner 无锡壹闪生物科技有限公司
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