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sTERM-1 enzyme-linked immunodetection kit and method thereof

An enzyme-linked immunoassay detection and kit technology, applied in the field of biomedicine, can solve the problem of low detection range and achieve the effects of large detection range, strong sensitivity and specificity, and high efficiency

Inactive Publication Date: 2012-08-15
GUANGZHOU GENESEED BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It is necessary to predict the TREM-1 content of the sample before the experiment, and the detection range is not high

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] Example 1: Preparation of sTERM-1 ELISA Kit

[0060] Proceed as follows:

[0061] (1) Coating: add 100ul 5ug / ml / s capture antibody to each well of the microtiter plate, and coat and react overnight at 4°C. The capture antibody is a capture antibody capable of antigen-antibody reaction with sTERM-1;

[0062] (2) Blocking: remove the coating buffer, add 100-200 μL bovine serum albumin solution to each well and wash 3 times, and the bovine serum albumin solution is a solution containing the following substances per liter:

[0063] NaHPO 4 .12H 2 O 3.72g,

[0064] K H 2 PO 4 0.43g,

[0065] NaCl 6.8g;

[0066] Then add 200-300 μL of blocking solution to each well to carry out blocking reaction for 2 hours. The blocking solution is a solution containing the following substances per liter:

[0067] BSA 10g,

[0068] 20g of sucrose,

[0069] Thimerosal 0.01g.

[0070] (3) Preservation: Shake off the liquid in the wells of the sealed microwell plate, pat ...

Embodiment 2

[0081] Embodiment 2: sample detection

[0082] (1) Add 100ul / well of the sample to be tested or sTREM-1 standard antigen on the microtiter plate coated with a capture antibody capable of antigen-antibody reaction with sTERM-1, and react at 37°C for 1 hour;

[0083] (2) Remove the reaction liquid in step 1, wash with washing buffer 3 times; add 200ng / ml HRP-labeled sTREM-1 detection antibody 50-200ul / well, react at 37°C for 30 minutes, the detection antibody can interact with sTERM-1 1 or antigen-antibody reaction with the combination of sTERM-1 and capture antibody after antigen-antibody reaction;

[0084] (3) Remove the sTREM-1 detection antibody solution; wash 5 times with washing buffer; add 100ul chromogenic substrate to each well and react at room temperature for 10 minutes, then add 1M HCL to each well to terminate the reaction, and read at a wavelength of 450nm on a microplate reader for analysis.

[0085] In a preferred embodiment of the present invention, the detecti...

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PUM

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Abstract

The invention discloses a sTERM-1 enzyme-linked immunodetection kit which comprises (1) an ELISA (enzyme-linked immunosorbent assay) plate and a detection antibody and (2) a reactant and a detection agent, wherein a capturing antibody capable of generating an antigen-antibody reaction with sTERM-1 covers and is fixed on the ELISA plate; the detection antibody can generate an antigen-antibody reaction with sTERM-1 or a combination obtained after the antigen-antibody reaction between the sTERM-1 and the capturing antibody; and the reactant and detection agent are used for detecting whether the sTERM-1 exists in a sample to be detected through enzyme-linked immunodetection. The scheme of the invention is simple in step and short in reaction time, reduces the difference of repeated experiments, is suitable for clinical detection and has great guiding significance to the prevention and diagnosis of multiple diseases, particularly acute pneumonia.

Description

technical field [0001] The invention relates to the technical field of biomedicine, in particular to a sTERM-1 enzyme-linked immunoassay kit, a preparation method of the kit and a method for detecting sTERM-1 using the kit. technical background [0002] TREM is a recently discovered immunoglobulin superfamily activating receptor expressed on myeloid cells. At least 5 members have been identified, among which TREM-1 is selectively expressed in neutrophils and monocytes, mediating The signal transduction pathway plays an important role in the occurrence and cascade amplification of the inflammatory response. [0003] Although the natural ligand of TREM-1 has not yet been identified, studies have found that activation of TREM-1 using a TREM-1 competitive monoclonal antibody can cause PMN degranulation, resulting in the release of pro-inflammatory factors and proteolytic enzymes, respiratory burst and cellular Phagocytosis, and this effect can synergize with Toll-like receptor ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577G01N33/558
Inventor 徐军发刘明
Owner GUANGZHOU GENESEED BIOTECH
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