Method for quantitative determination of specific analyte with single trapping agent and reagent kit therefor

An analyte and capture agent technology, applied in the field of bioengineering, can solve the problems of cumbersome detection methods, difficult protein labeling, inhibition of antibody binding, etc., and achieve the effects of high detection specificity, wide application range, and low noise

An analyte and capture agent technology, applied in the field of bioengineering, can solve the problems of cumbersome detection methods, difficult protein labeling, inhibition of antibody binding, etc., and achieve the effects of high detection specificity, wide application range, and low noise

CN1700009AInactive Publication Date: 2005-11-23孙东旭
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  • Method for quantitative determination of specific analyte with single trapping agent and reagent kit therefor
  • Method for quantitative determination of specific analyte with single trapping agent and reagent kit therefor
  • Method for quantitative determination of specific analyte with single trapping agent and reagent kit therefor

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Example 1: Singleness detection (detection of a protein)

[0036] In this embodiment, both the capture device and the detection device are 96-well microtiter plates. Each well of the capture device is coated with an antibody that detects an antigenic protein. Antigen proteins to be tested are four cytokines, namely IL-1-beta, IL-4, IL-8 and GM-CSF. The corresponding antibodies are all monoclonal antibodies.

[0037] (1) Capture antigenic protein:

[0038] a. Coated capture antibody: Take two 96-well microplates (flat bottom, high binding to protein), one for capturing antigen in the sample (capture device), and the other for detecting labeled antigen (detection device). 100 μl of monoclonal antibodies against IL-1-beta, IL-4, IL-8 or GM-CSF at a concentration of 0.5 μg / ml (diluted in PBS buffer) were added to the microwells. Only one antibody was added to each microwell, and each antibody was added to 8 microwells on each microplate. Place the microplate at 4 overn...

Embodiment 2

[0050] Embodiment 2: Multiplex detection (simultaneous detection of three proteins)

[0051] In this embodiment, both the capture device and the detection device are 96-well microtiter plates. The surface of each microwell of the capture device is coated with three kinds of antibodies mixed together to detect three antigenic proteins simultaneously. Antigen proteins to be tested were three cytokines, namely IL-1-beta, TNF-α and IL-10. The corresponding antibodies are all monoclonal antibodies.

[0052] (1) Capture antigenic protein:

[0053] a. Coating capture antibody: Take two 96-well microplates (flat bottom, high protein binding), one for capture (capture device) and one for detection (detection device). Add 100 μl of three antibody mixtures against IL-1-beta, TNF-α and IL-10 into the microwells of the capture device, the concentration of each antibody is 0.5 μg / ml (diluted in PBS buffer), add a total of 8 micropores. However, only one antibody is added to each microw...

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Abstract

The invention relates to a method and its medium box for quantity measuring peculiar analysis substance by single capture medium. The method comprises the following steps: combining the tested analysis substance with solid phase capture medium, labeling analysis substance which has been captured by capture medium with report molecule, cleaning-up analysis substance from compound, recombining tested analysis substance with new solid phase capture medium, ascertaining content of analysis substance by report molecule's label signal. The medium box comprises capture device, testing device report molecule used as label and analysis substance eluent.

Description

technical field [0001] The invention belongs to the technical field of bioengineering, and in particular relates to a method for quantitatively detecting specific analytes with a single capturing agent and a kit thereof. Background technique [0002] Detecting the content of specific protein factors in biological (including human) tissue samples is of great significance for applications and basic research in the fields of medicine, biology, agriculture, and the like. For example, the detection of specific protein components of pathogenic microorganisms is currently an important means of diagnosing infectious diseases; similarly, the determination of changes in the levels of early biomarker protein factors in cancer is extremely important for early detection and treatment of diseases and observation of curative effects. According to different application goals, the analyte to be tested can be one or several proteins, or a group of proteins with different numbers. In recent ye...

Claims

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Application Information

Patent Timeline
23 Nov 2005
Publication
CN1700009A
IPC
G01N33/543; G01N33/577
CPC
G01N33/54306
Inventors
孙东旭