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Biological sample detection method and detection kit

A detection kit and detection method technology are applied in the field of quantitative detection of biological samples, and can solve the problems such as the inability to achieve absolute quantification.

Pending Publication Date: 2020-10-27
RESUN (SHENZHEN) TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the Simoa method still relies on calculating the optical signal intensity of each reaction chamber to calculate the concentration through the Poisson formula, which cannot achieve complete absolute quantification.

Method used

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  • Biological sample detection method and detection kit
  • Biological sample detection method and detection kit

Examples

Experimental program
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Embodiment 1

[0078] This embodiment takes the detection of troponin cTnI as an example to further illustrate the detection method of the present invention.

[0079] 1. Preparation of antibody 1-labeled immunomagnetic beads

[0080] Immunomagnetic beads (Ademtech) were replaced by MES buffer three times, and the pH of the solution was 4-5, and the ionic strength was 0.1M. Add 1 mg each of EDC (1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride) and NHS (N-hydroxysuccinimide) dissolved in DMSO to the balanced solution , centrifuge and discard the supernatant after reacting for 20 minutes;

[0081] Reconstitute the precipitate after discarding the supernatant with the MES buffer, add capture troponin antibody 1 (a34600, biospacific), mix and coat for 1 hour, then add the terminator BSA (bovine serum albumin) to the concentration of BSA 1%, the reaction was terminated.

[0082] 2. Preparation of Antibody 2-labeled nanoparticles

[0083] 1 mg nanoparticles (polyethylene PS microsph...

Embodiment 2

[0098] This embodiment takes the simultaneous detection of troponin cTnI and troponin cTnT as an example to further illustrate the detection method of the present invention.

[0099] 1. Preparation of various antibody 1-labeled immunomagnetic beads

[0100] Immunomagnetic beads (Ademtech) were replaced by MES buffer three times, and the pH of the solution was 4-5, and the ionic strength was 0.1M. Add 1 mg each of EDC (1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride) and NHS (N-hydroxysuccinimide) dissolved in DMSO to the balanced solution , centrifuge and discard the supernatant after reacting for 20 minutes;

[0101] The precipitate after discarding the supernatant was redissolved with the MES buffer, and the capture troponin cTnI antibody 1 (a34600, biospacific) and the capture troponin cTnT antibody 1 (1C11, HyTest) were added, mixed and coated for 1 hour, and then The reaction was terminated by adding a terminator BSA (bovine serum albumin) until the concentrat...

Embodiment 3

[0110] This embodiment takes the simultaneous detection of troponin cTnI, troponin cTnT, C-reactive protein (CRP) and procalcitonin (PCT) as an example to further illustrate the detection method of the present invention.

[0111] 1. Preparation of various antibody 1-labeled immunomagnetic beads

[0112] Immunomagnetic beads (150 nm, Ademtech) were replaced by MES buffer three times, and the pH of the solution was 4-5, and the ionic strength was 0.1M. Add 1 mg each of EDC (1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride) and NHS (N-hydroxysuccinimide) dissolved in DMSO to the balanced solution , centrifuge and discard the supernatant after reacting for 20 minutes;

[0113] Reconstitute the precipitate after discarding the supernatant with the MES buffer, add cTnI antibody 1 (a34600, biopacific) to capture troponin, mix and coat for 1 hour, then add the terminator BSA (bovine serum albumin) to the concentration of BSA 1%, the reaction was terminated.

[0114] Immu...

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Abstract

The invention discloses a biological sample detection method and a detection kit. The method comprises the following steps: forming a ternary complex (magnetic bead-antigen-marker) in a manner of immunospecific binding with magnetic beads, and collecting the ternary complex under the action of a magnetic field; under a specific chemical reagent, eluting a markers in the ternary immune complex, separating the markers from magnetic beads under the action of the magnetic field, and collecting eluent containing the markers, and re-dispersing the eluent by using strong electrolyte; and detecting the eluent containing the markers and then, realizing absolute quantification through a nano-particle counter. The method disclosed by the invention can be used for detecting trace protein below the lower limit of conventional immunodetection, and can be widely applied to the fields of immunological detection, microbiological detection, cell separation and the like.

Description

technical field [0001] The invention belongs to the technical field of biomolecular detection, and in particular relates to a quantitative detection method of biological samples. Background technique [0002] The occurrence and development of diseases are closely related to the abnormal expression of proteins or the expression of specific proteins. Accurate determination of the content of disease-related proteins is of great significance in the prevention and control of infectious diseases, cancer screening and accurate diagnosis. [0003] Several commonly used immunological techniques, enzyme-linked immunoassay is currently the most widely used immunoassay method. This method is to combine the specificity of the antigen-antibody reaction with the enzyme-catalyzed substrate on the secondary antibody, and judge the test result according to the color change after the enzyme acts on the substrate, and its sensitivity can reach the ng level. Commonly used enzymes for labeling ...

Claims

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Application Information

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IPC IPC(8): G01N33/537G01N33/543G01N33/68G01N33/74G01N33/58G01N15/10
CPCG01N33/5375G01N33/54326G01N33/54346G01N33/6854G01N33/74G01N33/582G01N33/587G01N15/10G01N2333/4712G01N2333/4737G01N2333/585
Inventor 王哲柳可熊贵
Owner RESUN (SHENZHEN) TECH CO LTD
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