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

An electrochemical and reagent kit technology, applied in the field of immunoassay, can solve the problems such as the absence of high-sensitivity and high-specificity quantitative detection methods or reagent kits for AChE-Ab, achieve fast detection speed, rapid detection, and avoid result deviations Effect

Pending Publication Date: 2020-04-17
江苏三联生物工程股份有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] The technical problem to be solved in the present invention is: in order to solve the technical problem that there is no method or kit for quantitatively detecting AChE-Ab with high sensitivity and high specificity in the prior art, a kind of electrochemiluminescent reagent for detecting AChE-Ab is provided box and method

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

Examples

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Effect test

Embodiment 1

[0020] Embodiment 1 detects the preparation method of the electrochemiluminescence kit of AChE-Ab

[0021] 1) Preparation of working solution of magnetic beads coated with streptavidin

[0022] The suspension of streptavidin-coated magnetic beads was magnetically separated to remove the supernatant, and resuspended to a concentration of 0.75 mg in PBS buffer (0.1 M, pH 7.4) with a concentration of 0.1 mol / 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.

[0023] 2) Preparation of biotin-labeled AChE antigen

[0024] Accurately weigh 1 mg of AChE 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. Once, transfer the antibody to a centrifuge tube or a cryovial after dialysis;

[0025] Accurately weigh 1mg of biotin activated by N-hydroxys...

Embodiment 2

[0041] Example 2 The method of using the kit of the present invention to detect AChE-Ab uses a fully automatic electrochemiluminescence immunoassay analyzer as a detection instrument, and the kit is loaded on the instrument for detection, and the steps are as follows:

[0042] Add the sample (or calibrator or quality control), biotin-labeled AChE antigen working solution and ruthenium-tripypyridyl-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;

[0043] 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;

[0044] The magnetic composite suspension is placed in a magnetic field, and the cleaning solution flows through and washes the magnetic composite;

[0045] Inject the electrochemiluminescent substrate solution into the washed magneti...

Embodiment 3

[0046] The performance evaluation of embodiment 3 kits of the present invention

[0047] 1) Research on reagent sensitivity

[0048] 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 mean 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 results between the calibrator (0.2ng / mL) were subjected to two-point regression fitting to obtain a linear equation, and the RLU value corresponding to M+2SD was brought into the above equation to calculate the corresponding concentration, namely LOB.

[0049] The measurement results in Table 1 below show that the LOB of the reagent detected by th...

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Abstract

The invention relates to a novel electrochemiluminescence kit for detecting an acetylcholin esterase antibody (AChE-Ab) from blood and a preparation method of the novel electrochemiluminescence kit. The prepared kit comprises a streptavidin-coated magnetic bead particle working solution, a biotin-labeled AChE antigen working solution, a terpyridyl ruthenium-labeled anti-human IgG antibody workingsolution, an AChE-Ab calibration material and / or a quality control material working solution, a tripropylamine-containing electrochemiluminescence substrate solution and a cleaning solution. A luminescence system of the prepared kit 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, the detection result repeatability is good, and accurate quantitative detection of the AChE-Ab in the blood can be achieved.

Description

technical field [0001] The invention relates to an electrochemiluminescence reagent kit for detecting AChE-Ab and its usage, belonging to the technical field of immune analysis. Background technique [0002] Myasthenia gravis (MG) is an autoimmune disease, which is an autoantibody-mediated, complement-involved, cellular immune-dependent neurotransmitter signal transmission disorder at the neuromuscular junction (NMJ). The incidence rate of MG population is (32-64) / 100,000, and there are about 600,000 MG patients in my country [1]. The main clinical manifestations are skeletal muscle weakness and fatigue, which aggravate after exercise, and the symptoms are obviously relieved and alleviated after rest and application of cholinesterase inhibitors. It is characterized by high morbidity, high disability rate, poor prognosis, and easy recurrence, which has seriously threatened human life and health, and has received extensive attention from medical and scientific researchers. M...

Claims

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Application Information

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IPC IPC(8): G01N33/543G01N27/26G01N21/76
CPCG01N21/76G01N27/26G01N33/54326G01N2800/24
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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