Electrochemical luminescence kit for detecting RyR-Ab and preparation method thereof

An electrochemical and reagent kit technology, applied in the field of immunoassay, to achieve fast detection, avoid result deviation, and improve work efficiency

Pending Publication Date: 2020-04-14
江苏三联生物工程股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

There is currently no electrochemiluminesc

Method used

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  • Electrochemical luminescence kit for detecting RyR-Ab and preparation method thereof
  • Electrochemical luminescence kit for detecting RyR-Ab and preparation method thereof
  • Electrochemical luminescence kit for detecting RyR-Ab and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Example 1 The preparation method of the electrochemiluminescence kit for detecting RyR-Ab

[0022] 1) Preparation of working solution of magnetic beads coated with streptavidin

[0023] The suspension of streptavidin-coated magnetic beads was magnetically separated to remove the supernatant, and resuspended to a concentration of 0.75mg with PBS buffer (0.1M, pH 7.4) with a concentration of 0.1mol / L at pH=7.4 / mL, the buffer contains 0.5 wt% BSA, 0.05 wt% Triton X-100, 0.05 wt% proclin 300 and 0.05 wt% sodium azide.

[0024] 2) Preparation of biotin-labeled RyR antigen

[0025]Accurately weigh 1 mg of RyR antigen, add an appropriate amount of PBS buffer (0.1M, pH 7.4) to adjust the total antibody concentration to 1 mg / mL, and add it to a dialysis bag for dialysis. Change the dialysate every 3 to 4 hours for 3 to 4 hours. 4 times, transfer the antibody to a centrifuge tube or cryopreservation tube after dialysis;

[0026] Accurately weigh 1mg of biotin activated by N-h...

Embodiment 2

[0042] Example 2 The method of using the kit of the present invention to detect RyR-Ab uses a fully automatic electrochemiluminescence immunoassay analyzer as a detection instrument, and loads the kit on the instrument for detection. The steps are as follows:

[0043] Add the sample (or calibrator or quality control), biotin-labeled RyR antigen working solution and ruthenium-tripyridyl-labeled anti-human IgG antibody working solution into the cuvette, and incubate at 37°C for 10 minutes to form antigen-antibody-antibody Antibody complex solution;

[0044] Add streptavidin-coated magnetic bead working solution to the antigen-antibody-antibody complex solution, and incubate at 37°C for 10 minutes to form a magnetic complex suspension;

[0045] The magnetic composite suspension is placed in a magnetic field, and the cleaning solution flows through and washes the magnetic composite;

[0046] Inject the electrochemiluminescent substrate solution into the washed magnetic complex, a...

Embodiment 3

[0047] The performance evaluation of embodiment 3 kits of the present invention

[0048] 1) Research on reagent sensitivity

[0049] Reagent sensitivity is determined according to the minimum detection limit (Limit of Blank, LoB), and the minimum detection limit is carried out according to the following experimental method. Detect the zero-concentration calibrator 20 times, get the signal value (RLU) of 20 measurement results, calculate the average value M and standard deviation SD, and obtain the RLU value corresponding to M+2SD, according to the zero-concentration calibrator and the adjacent concentration The concentration-RLU mean value between the calibrator (0.2ng / mL) is obtained by two-point regression fitting to obtain a linear equation, and the RLU value corresponding to M+2SD is brought into the above equation to calculate the corresponding concentration, that is LoB.

[0050] The measurement results in Table 1 below show that the LoB of the reagent detected by the ...

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Abstract

The invention relates to a novel electrochemical luminescence kit for detecting an anti-Ryanodine receptor antibody (RyR) from a blood sample and a preparation method thereof. The prepared kit comprises a streptavidin-coated magnetic bead particle working solution, a biotin-labeled RyR antigen working solution, a terpyridyl-ruthenium-labeled anti-human IgG antibody working solution, a RyR-Ab calibration material and/or quality control material working solution, a tripropylamine-containing electrochemiluminescence substrate solution and a cleaning solution. A luminescence system of the preparedkit is electrochemiluminescence, a streptavidin-biotin biological reaction signal amplification system is utilized, the detection speed is high, the sensitivity is high, the linear range is wide, thedetection result repeatability is good; and accurate quantitative detection of RyR-Ab in blood can be realized.

Description

technical field [0001] The invention relates to an electrochemiluminescence reagent kit for detecting RyR-Ab and its usage, belonging to the technical field of immune analysis. Background technique [0002] Autoimmune disease (hereinafter referred to as "autoimmunity") is the immune system's immune response to the components of the body, causing damage and causing disease. Under normal circumstances, the immune system only responds to foreign substances that invade the body, such as bacteria, viruses, parasites, and transplants, and destroys or rejects these foreign substances. Under the influence of certain factors, certain abnormalities appear in the tissue components of the body or the immune system itself, causing the immune system to mistake its own components as foreign objects and attack them. At this time, the immune system will produce antibodies against some of the body's own components and active lymphocytes, which will damage and destroy its own tissues and orga...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/531G01N27/26G01N21/76
CPCG01N21/76G01N27/26G01N33/531G01N33/6872
Inventor 卜梅杰泮锋纲丁俊杰施启尧
Owner 江苏三联生物工程股份有限公司
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