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Kit for in vitro detection of GPI (Glucose-6-Phosphate Isomerase) antigen and preparation method thereof

A phosphoisomerase and antigenic technology, applied in the field of bioengineering, can solve the problems of long time and complicated procedures for detecting GPI concentration, and achieve the effects of convenient detection results, accurate detection results and significant technological progress.

Active Publication Date: 2012-01-18
上海北加生化试剂有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to provide a glucose-6-phosphate isomerase antigen in vitro detection kit and its preparation method. The glucose-6-phosphate isomerase antigen in vitro detection kit and its preparation method adopt colloidal gold The principles of diafiltration and chromatography are used to detect the concentration of GPI in human serum, which solves the technical problems of long time and complicated procedures in the prior art for detecting the concentration of GPI in serum

Method used

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  • Kit for in vitro detection of GPI (Glucose-6-Phosphate Isomerase) antigen and preparation method thereof
  • Kit for in vitro detection of GPI (Glucose-6-Phosphate Isomerase) antigen and preparation method thereof
  • Kit for in vitro detection of GPI (Glucose-6-Phosphate Isomerase) antigen and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Embodiment 1. colloidal gold preparation condition selection:

[0019] 1. 300 ml of 10.05% gold chloride solution, heated to boiling;

[0020] 1.2 Add 3.6 ml of 5.0% trisodium citrate;

[0021] 1.3 Continue heating and boiling for 15 minutes, stop heating, wait until it cools to room temperature, and restore to the original volume with distilled water;

[0022] 1.4 Experimental results:

[0023] Wavelength λ: 540 535 530 525 520 510nm.

[0024] ABS value: 0.528 0.715 1.481 0.902 0.699 0.601

[0025] Visually observe the color of the colloidal gold solution: red and transparent.

Embodiment 2

[0026] Embodiment 2. Colloidal gold conjugate preparation condition selection:

[0027] 2.1 Selection of pH value of colloidal gold-labeled goat anti-GPI antibody:

[0028] Select the appropriate pH value of the colloidal gold-labeled goat anti-GPI antibody, use 1.0% anhydrous sodium carbonate solution to adjust the pH value of the colloidal gold-labeled antibody, and compare the appropriate pH range of the colloidal gold-labeled goat anti-GPI antibody through detection sensitivity.

[0029] Experimental results:

[0030]

[0031] The experimental results showed that the pH range of the colloidal gold markers had better sensitivity in the pH7.2-7.5 and pH7.5-8.0 groups, but when the positive spots were observed visually, the positive spots in the pH7.5-8.0 group were found to be darker and darker. Therefore, it is recommended that the pH range of the colloidal gold label should be controlled at pH 7.2-7.5, that is, for every 100 ml of colloidal gold, add 2.4 ml of 1.0% anh...

Embodiment 3

[0036] Embodiment 3. Detection plate preparation condition selection:

[0037] 3.1 Comparison of the pH conditions of the coating medium at the detection point:

[0038] Experimental Materials:

[0039] (1) Nitrocellulose membrane with a pore size of 0.45 μm;

[0040] (2) Coating buffer: 1.0M NaHCO 3 ;

[0041] NaHCO 3 (Analytical pure) 8.401 grams, put into container, measure process water with graduated cylinder and pour into container, wait to dissolve, then take process water to make up volume to 100 milliliters, mix;

[0042] (3) Detection point antibody: goat anti-GPI antibody (1.0mg / mL);

[0043] experiment procedure:

[0044] (1) Detection point antigen processing:

[0045] Goat anti-GPI antibody (1.0mg / mL) stock solution, pH=7.0-7.5, neutral;

[0046] Goat anti-GPI antibody (1.0mg / mL) stock solution 0.05ml add 1.0M NaHCO 3

[0047] 0.005ml, mix well, pH≥8.0;

[0048] (2) Coating: Take 2.0uL of the detection point antigen and add it on the nitrocellulose mem...

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Abstract

The invention belongs to the field of biological engineering, and provides a kit for in vitro detection of a GPI (Glucose-6-Phosphate Isomerase) antigen, which comprises a detection plate, a goat anti-GPI antibody, a colloidal gold conjugate, a confining liquid, a washing liquid, a positive reference product and a negative reference product. The invention also provides a preparation method of the kit for the in vitro detection of the GPI antigen. The kit adopts a double antibody sandwich colloidal gold immunosorbent assay method principle to detect the GPI antigen in a human serum, the goat anti-GPI antibody is coated on a nitrocellulose membrane to be made into a solid phase antibody for capturing the GPI antigen which possibly exists in a human peripheral serum, and then colloidal gold is labeled on the goat anti-GPI antibody to form the colloidal gold conjugate as a tracer; and if the detected human serum contains the GPI antigen, then a solid phase goat anti-GPI antibody-GPI antigen-colloid gold conjugate is formed. The invention can be applied to the auxiliary diagnosis of rheumatoid arthritis.

Description

technical field [0001] In the field of bioengineering, the present invention particularly relates to a kit and a preparation method thereof, in particular to a glucose-6-phosphate isomerase antigen in vitro detection kit and a preparation method thereof. Background technique [0002] Rheumatoid arthritis (RA) is the most common autoimmune disease, affecting approximately 1.0% of the world's population. It is characterized by symmetrical inflammation of the synovial membranes of the large and small joints. Initial symptoms include swelling and pain in the joints, and morning stiffness in the joints, which can lead to bone destruction. Therefore, early diagnosis of RA and timely initiation of appropriate treatment are very important to control the condition of the disease. The number of patients with rheumatoid arthritis (RA) in small and medium-sized hospitals is small, the number of test samples is small, and the test report cycle is long, which easily delays the diagnosis...

Claims

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Application Information

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IPC IPC(8): G01N33/544G01N33/532
Inventor 刘颖冰陆梅生刘中令
Owner 上海北加生化试剂有限公司
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