Nerve specificity olefinic alcohol enzyme chemiluminescence immune analysis determination reagent kit and preparing method thereof

A kind of enolase chemistry and enolase technology, which is applied in the field of immunoanalysis medicine, can solve the problems of expensive instruments, limited popularization and use, unstable measurement results, etc., and achieve the effect of improving sensitivity

Active Publication Date: 2009-03-04
CHEMCLIN DIAGNOSTICS CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Radioimmunoassay has many disadvantages, such as complex operation, unstable measurement results, short storage time of reagents, radioactive contamination, expensive instruments, etc.
Enzyme immunoassay has low sensitivity and many influencing factors, which can easily cause false negatives and false positives
[0005] The chemiluminescence immunoassay quantitative kit in the prior art is a closed automatic chemiluminescence measurement system, which requires an expensive fully automatic chemiluminescence measuring instrument, thus limiting its popularization and use, and cannot be widely used in clinical diagnosis and scientific research

Method used

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  • Nerve specificity olefinic alcohol enzyme chemiluminescence immune analysis determination reagent kit and preparing method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Example 1 Preparation of Quantitative Assay Kit for Nerve-Specific Enolase Chemiluminescent Immunoassay

[0048] 1. Preparation of enzyme-labeled antibody

[0049] (1) Preparation of horseradish peroxidase-labeled nerve-specific enolase monoclonal antibody

[0050] Dissolve 4.4mg HRP in 1mL distilled water, add 0.4mL sodium periodate (50mmol / L) and stir at room temperature for 20min, dialyze through 1mmol / L sodium acetate buffer solution, pH 4.4, add 8mg nerve-specific enolase antibody, stir 2h, finally use 200mmol / L NaBH 4 For reduction, after dialysis with 0.02M PB buffer, add an equal volume of glycerol and store below -20°C.

[0051] (2) Preparation of alkaline phosphatase-labeled nerve-specific enolase monoclonal antibody

[0052] Nerve-specific enolase monoclonal antibody was coupled with alkaline phosphatase by the glutaraldehyde method, fully dialyzed against PBS, added an equal volume of glycerol, and stored below -20°C.

[0053] 2. Preparation of solid-pha...

Embodiment 2～3

[0107] Examples 2-3 Preparation of the nerve-specific enolase chemiluminescent immunoassay quantitative assay kit of the present invention

[0108] The quantitative assay kit of the present invention was prepared in the same manner as in Example 1 except that plastic beads and magnetic particles were used as carriers respectively.

Embodiment 4

[0109] Embodiment 4 The usage method of the kit of the present invention

[0110] The specific operations of the kit prepared in the above embodiment 1 are as follows:

[0111] 1) Equilibrate all detection reagents and samples to room temperature;

[0112] 2) Take the required amount of slats;

[0113] 3) Add 50 μL of enzyme-labeled antibody, calibrators / sample to be tested in turn to each well and tube, shake on a micro shaker for 30 seconds to mix evenly;

[0114] 4) Incubate at room temperature for 60 minutes;

[0115] 5) wash the plate with an automatic enzyme-labeled plate washer, and separate the coated antibody-antigen-enzyme-labeled antibody complex immobilized on the solid phase carrier from unbound substances;

[0116] 6) Mix luminescent substrates A and B at a ratio of 1:1, add a volume of 100 μL to each well / tube, incubate at room temperature for 5 minutes, and use a chemiluminescence detector for detection within 5 to 15 minutes;

[0117] 7) Use the double log...

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Abstract

The invention relates to the medical field of immunoassay, more specially, the invention provides a chemiluminescent immunoassay detection kit for nerve specific enolase and a preparation method thereof. The kit of the invention comprises 1) nerve specific enolase calibrators, 2) solid-phase vectors which are coated by nerve specific enolase monoclonal antibodies, 3) nerve specific enolase monoclonal antibody antigens which are marked by enzyme and 4) chemiluminescent substrates which are acted by the enzyme. The invention also provides a preparation method of the kit, which comprises the steps: preparing the calibrators, coating the vectors, marking the antibodies, packaging, and the like. The kit of the invention has the advantages of convenience, rapidness, sensitivity, stability, and the like. The invention can provide good technical support for the early detection, the detection of disease progress and the judgment of healing effect of some diseases such as small cell lung carcinoma (SCLC), neuroblastoma, brain damage, and the like.

Description

technical field [0001] The invention relates to the field of immunoanalysis medicine, in particular, the invention provides a quantitative assay kit for nerve-specific enolase chemiluminescence immunoassay and a preparation method thereof. Background technique [0002] Enolase is a key enzyme in the glycolysis process, which is composed of dimers composed of three subunits α, β and γ, which can be divided into five isozymes (αα, αβ, αγ, ββ and γγ). Among them, the γ-dimer isoenzyme NSE (γγ, nerve-specific enolase) was named after it was first discovered to exist in nerve cells and neuroendocrine cells. In addition to the common catalytic activity of enzymes, NSE is also related to the differentiation and maturation of nerve and neuroendocrine cells. In recent years, NSE has attracted increasing attention as a sensitive tumor marker and a quantitative indicator of brain injury. [0003] In the chemical classification of tumor markers, NSE belongs to enzyme markers. NSE is...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577G01N33/543G01N21/76
Inventor 林珍林金明王栩李振甲应希堂胡国茂唐宝军
Owner CHEMCLIN DIAGNOSTICS CO LTD
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