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Diluted composition and reagent or kit for osteopontin detection

A technology for osteopontin and detection reagents, which is applied in the field of immunological detection to achieve the effects of saving time, shortening detection time, and eliminating the difference between old and new samples

Inactive Publication Date: 2020-06-19
AUTOBIO DIAGNOSTICS CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] At present, the quantitative detection method of OPN in the market is mainly enzyme-linked immunoassay (Wuhan Boster, American AssayDesigns, Beijing Haitai Tongda), which has certain limitations in detection accuracy, linear range, time-consuming, safety and operation. sex

Method used

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  • Diluted composition and reagent or kit for osteopontin detection
  • Diluted composition and reagent or kit for osteopontin detection
  • Diluted composition and reagent or kit for osteopontin detection

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preparation example Construction

[0070] The preparation method of described calibrator is: calibrator diluent comprises the PBS buffer solution of PH=7.4, preservative is 1‰ (w / v) Proclin 300 and 1‰ (w / v) Bro; Protein stabilizer is 1 % (w / v) casein; 0.2‰ of sunset yellow or tartrazine; dilute the OPN antigen into 6 gradients of 0ng / mL, 5ng / mL, 10ng / mL, 50ng / mL, 100ng / mL with calibrator diluent mL, 200ng / mL, respectively recorded as S0, S1, S2, S3, S4, S5, divided into 1ml / bottle, stored at 2-8°C.

[0071] The kit of the present invention adopts the method of magnetic particle chemiluminescence, and is matched with the AutoLumo A2000 series automatic chemiluminescence measuring instrument to realize full automation; it is convenient to use and simple to operate; the detection time of the kit is short, which greatly saves the time of inspectors ( The result is obtained in about 40 minutes); the kit of the present invention adopts the double-antibody sandwich method, which has a wide linear range and strong clin...

Embodiment 1

[0074] The preparation of embodiment 1 reagent

[0075] (1) Preparation of sealing solution

[0076] The sealing solution is prepared by Tris buffer solution with pH=7.4, and the preservatives contained therein are 1‰ (w / v) Proclin300 and 1‰ (w / v) NaN3; the protein stabilizer contained is 1% (w / v) v) bovine serum albumin; additionally 5% (w / v) glycerol was added.

[0077] (2) Preparation of enzyme conjugate dilution

[0078] The enzyme conjugate is made of Tris buffer at pH=7.4, containing 1‰ (w / v) Proclin 300 and 1‰ (w / v) Bro as preservatives; 25% (w / v) protein stabilizer Bovine serum; surfactant 1‰ (w / v) Triton X-100; 0.2‰ carmine pigment, stored at 2-8°C until use.

[0079] (3) Preparation of sample diluent

[0080] The sample diluent is PBS buffer solution with PH=7.4, containing 1‰ (w / v) Proclin 300 and 1‰ (w / v) Bro as preservatives; 1% (w / v) protein stabilizer Casein; 1% (w / v) disodium edetate, 0.2‰ chlorophyll, stored at 2-8°C until use.

[0081] (4) Preparation o...

Embodiment 2

[0095] The detection step of embodiment 2 kit of the present invention

[0096] 1) Into the reaction container (hereinafter referred to as "well"), sequentially add the calibrator (for calibration) and the sample, and the sample volume is 25 μL / well.

[0097] 2) Add 20 μL of magnetic particle suspension and 50 μL of sample diluent to each well.

[0098] 3) After mixing, incubate at 37°C for 15 minutes.

[0099] 4) Wash with cleaning solution 5 times.

[0100] 5) Add 100 μL of enzyme conjugate to each well.

[0101] 6) After mixing, incubate at 37°C for 17 minutes.

[0102] 7) Wash with cleaning solution 5 times.

[0103] 8) Add 50 μL each of substrate A solution and substrate B solution to each well.

[0104] 9) Check the luminescence value within 1 to 5 minutes after mixing.

[0105] 10) Result calculation

[0106] Select the appropriate curve fitting method to calculate the result. This kit recommends a four-parameter fitting method, with the concentration value of t...

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Abstract

The invention relates to the field of immunological detection, in particular to a diluted composition and a reagent or a kit for osteopontin detection. The kit for detecting osteopontin by a magneticparticle chemiluminescence method comprises a magnetic particle suspension coated with an osteopontin monoclonal antibody, an enzyme conjugate containing the osteopontin monoclonal antibody, a samplediluent and a calibrator. The kit is prepared based on the magnetic particle chemiluminiscence method, is used in cooperation with an Auto Lumo series full-automatic immunoassay instrument, can achieve automatic detection, and is easy and convenient to operate and high in detection speed. The kit has the advantages of wide detection range, high sensitivity and strong specificity, can quantitatively detect the content of osteopontin in serum and plasma, and is used for early diagnosis, disease process monitoring, treatment effect evaluation and prognosis monitoring of gastric cancer, lung cancer, liver cancer, breast cancer and the like.

Description

technical field [0001] The invention relates to the field of immunological detection, in particular to a dilution composition and a reagent or kit for detecting osteopontin. Background technique [0002] Osteopontin (OPN) is an endocrine non-collagenous glycosylated phosphoprotein that can be expressed by various cells such as osteoclasts, vascular smooth muscle cells, vascular endothelial cells, and macrophages. As a new type of cytokine, osteopontin is involved in bone metabolism, tumor growth and metastasis, inflammation and immunity. OPN can promote tumor cell chemotaxis, adhesion and migration, and can also promote the degradation of extracellular matrix. The expression is enhanced in pathological conditions (immune diseases, inflammation and tumors), such as liver cancer, lung cancer, gastric cancer, thyroid cancer, breast cancer, ovarian cancer, prostate cancer, and skin cancer. It can also be expressed at a high level. Studies have shown that OPN can promote tumor ...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/535G01N33/543G01N21/76
CPCG01N21/76G01N33/535G01N33/54326G01N33/6863
Inventor 吴小田崔小妞史小芹付光宇吴学炜
Owner AUTOBIO DIAGNOSTICS CO LTD
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