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Test strip for quantitatively detecting calprotectin, preparation method thereof and determining method for calprotectin concentration

A calprotectin, quantitative detection technology, applied in the biological field, can solve the problems of long detection time, instability, low sensitivity, etc., and achieve the effect of being suitable for professional department operations, shortening detection time, and convenient clinical use.

Inactive Publication Date: 2016-07-20
付国亮
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The ELISA method is widely used in hospital laboratories due to its quantitative accuracy, but the detection time is long and inconvenient; although the colloidal gold method is convenient and fast, it is a qualitative method with low sensitivity; the connection method between colloidal gold particles and antibodies is an electrostatic method. Adsorption, falling off during the detection process may cause inaccurate and unstable results

Method used

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  • Test strip for quantitatively detecting calprotectin, preparation method thereof and determining method for calprotectin concentration
  • Test strip for quantitatively detecting calprotectin, preparation method thereof and determining method for calprotectin concentration
  • Test strip for quantitatively detecting calprotectin, preparation method thereof and determining method for calprotectin concentration

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preparation example Construction

[0041] The preparation method of the above-mentioned test strip comprises the following steps: sequentially fixing a sample absorbent pad, a labeled antibody pad, a coating film and a water-absorbent pad on the bottom plate of the test strip;

[0042] Described coated film is prepared by following method:

[0043] Use the first phosphate buffer to dilute the rabbit anti-mouse IgG and calprotectin antibody respectively to obtain the rabbit anti-mouse IgG solution and the calprotectin antibody solution, the concentration of the first phosphate buffer is 0.01-0.03M, pH Value is 7.2-7.6, and the concentration of described rabbit anti-mouse IgG solution and calprotectin antibody solution is 1mg / ml-1.5mg / ml; Spray it on the nitrocellulose membrane at an interval of 0.5cm-1.0cm, dry it at 35-38℃ for 0.5-1.5h, add a desiccant and seal it for later use.

[0044] The labeled antibody pad is prepared by the following method:

[0045] Prepare the calprotectin antibody solution: dialyze ...

Embodiment 1

[0050] Preparation of fluorescent immunochromatographic test strips for calprotectin:

[0051] A. Preparation of antibody: select commercialized calprotectin monoclonal antibody, dialyze overnight (18h) at 4°C with 0.05MpH8.5 borate buffer solution; select commercialized calprotectin monoclonal antibody, use 0.02MpH7 .4 PBS 4 ℃ dialyzed overnight (16h);

[0052] B, the preparation of the fluorescent microsphere cushion of labeling calprotectin monoclonal antibody: select the fluorescent microsphere (Bangslab company, excitation wavelength 350nm, detection wavelength 615nm) that diameter is 100nm for use, adjust microsphere with 0.05M, pH4.5MES buffer The concentration is 1% by mass fraction, and then the calprotectin monoclonal antibody is labeled onto the fluorescent microspheres by means of covalent coupling of carbodiimide (EDC) and succinimide (NHS). Spray the prepared fluorescent microspheres on the fluorescent microsphere pad at an amount of 3 μl / cm using a quantitative...

Embodiment 2

[0062] The test strip preparation method of the present embodiment comprises the following steps:

[0063] A. Preparation of antibody: select commercialized calprotectin monoclonal antibody, dialyze overnight at 4°C with 0.05MpH8.5 borate buffer solution; select commercialized calprotectin monoclonal antibody, use 0.02MpH7.4 borate buffer Dialyze overnight at 4°C in PBS;

[0064] B. Preparation of colloidal gold pad labeled with calprotectin monoclonal antibody:

[0065] Take 50ml of colloidal gold and filter.

[0066] Dilute the antibody 3 times with water (about 1 mg / ml), add it within 30 minutes, and mix well while adding it. The mixing force should be small near the ice to prevent damage to the connection between the antibody and the colloidal gold.

[0067] 10% BSA was added to make the final concentration of the solution 1%, and 10% PEG was added to make the final concentration 0.2%, and the addition was completed within 15 minutes.

[0068] Centrifuge at 10000r / min f...

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Abstract

The invention discloses a test strip for quantitatively determining calprotectin, a preparation method thereof and a determining method for calprotectin concentration, belonging to the technical field of biotechnology. The test strip comprises a base plate, wherein a sample absorbing pad, a labeled antibody pad, a wrapped film and a water absorbing pad are successively arrayed and attached to the base plate. The test strip provided by the invention has the advantages of accurate test result, convenience and low cost.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to an immunochromatographic test strip for quantitatively detecting human calprotectin (calprotectin), a preparation method thereof, and an instant and rapid determination method for the concentration of calprotectin (calprotectin) . Background technique [0002] Calprotectin is a calcium and zinc binding protein with a molecular weight of 36kD. It consists of two heavy chains with a molecular weight of 14kD and a light chain with a molecular weight of 8kD, which are covalently linked calcium-binding protein heterotrimers. [0003] Calprotectin is considered to be a reliable indicator of inflammation in many diseases, and it can clearly distinguish organic diseases such as inflammatory diseases such as IBD from functional diseases such as IBS. This in vitro diagnostic tool can avoid the costly and invasive procedure of colonoscopy. The annual meeting of the British Society of Gastroe...

Claims

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Application Information

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IPC IPC(8): G01N33/68
CPCG01N33/68
Inventor 付国亮段学军
Owner 付国亮
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