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Colloidal gold labeling method for improving sensitivity and application and product thereof

A labeling method, colloidal gold technology, applied in the field of immunochromatography, can solve the problems of low overall efficiency, easy cross-linking dead gold, strong electrostatic attraction, etc., to achieve the effect of protecting protein activity, weakening electrostatic attraction, and increasing viscosity

Pending Publication Date: 2022-05-13
SHANGHAI BIOGERM MEDICAL TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although this method improves the labeling efficiency to a certain extent, the labeling needs to be cooled and loaded slowly for re-alkali, and the operation is relatively complicated; and the colloidal gold-protein conjugate before re-alkali has strong electrostatic attraction in an acidic environment, and is easy to Cross-linked dead gold, which leads to the aggregation of colloidal gold, even after re-alkali, the aggregated conjugates cannot be re-dispersed and re-dissolved, and the overall efficiency is not high

Method used

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  • Colloidal gold labeling method for improving sensitivity and application and product thereof
  • Colloidal gold labeling method for improving sensitivity and application and product thereof
  • Colloidal gold labeling method for improving sensitivity and application and product thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0068] The determination of embodiment 1 optimum labeling pH

[0069] Taking 2019-nCoV N-antibody-1 as an example, the isoelectric point of 2019-nCoV N-antibody-1 is 7.9, and the optimal labeling pH of 2019-nCoV N-antibody-1 treated with different protein activators was compared.

[0070] Protein Activator 1: Purified Water.

[0071] Protein activator 2: ON-870 with a final mass concentration of 0.005%, trehalose with a final mass concentration of 0.2%, tartaric acid with a final mass concentration of 0.1%, and purified water.

[0072] Protein activator 3: ON-870 with a final mass concentration of 0.05%, trehalose with a final mass concentration of 0.2%, tartaric acid with a final mass concentration of 0.1%, and purified water.

[0073] Protein activator 4: ON-870 with a final mass concentration of 0.01%, trehalose with a final mass concentration of 0.2%, tartaric acid with a final mass concentration of 0.1%, and purified water.

[0074] Protein activator 5: ON-870 with a fi...

Embodiment 2

[0085] Example 2 Preparation of novel coronavirus antigen detection kit

[0086] 1. Preparation of gold-labeled conjugates

[0087] Take 10mL colloidal gold to the beaker, add 0.1mol / L K 2 CO 3 Adjust the pH of the colloidal gold solution to 0.3-0.8 lower than the isoelectric point of the novel coronavirus N antibody-1, and stir at 250r / min for 5 minutes;

[0088] Take 80 μg of novel coronavirus N antibody-1, add protein activator, dilute novel coronavirus N antibody-1 to 1.0mg / mL, mix well and let stand for 10 minutes;

[0089] Add the processed novel coronavirus N antibody-1 to the colloidal gold and stir for 30 minutes;

[0090] Add 200 μL 10% bovine serum albumin and continue stirring for 15 minutes;

[0091] Centrifuge at 7500r / min at 4°C for 35min, discard the supernatant, measure the absorbance with a UV spectrophotometer and calculate the OD value and labeling efficiency.

[0092] The protein activator comprises the following components: ON-870 with a final mass c...

Embodiment 3

[0104] The preparation of embodiment 3 adenovirus IgM antibody detection kit

[0105] 1. Preparation of gold-labeled conjugates

[0106] Take 10mL colloidal gold to the beaker, add 0.1mol / L K 2 CO 3 Adjust the pH of the colloidal gold solution to 0.3-0.8 lower than the isoelectric point of the adenovirus antigen, and stir at 250r / min for 5 minutes;

[0107] Take 80 μg of adenovirus antigen, add protein activator, dilute the adenovirus antigen to 1.mg / mL, mix well and let stand for 10 minutes;

[0108] Add the processed adenovirus antigen to the colloidal gold and stir for 30 minutes;

[0109] Add 200 μL 10% bovine serum albumin and continue stirring for 15 minutes;

[0110] Centrifuge at 7500r / min at 4°C for 35min, discard the supernatant, measure the absorbance with a UV spectrophotometer and calculate the OD value and labeling efficiency.

[0111] The protein activator comprises the following components: ON-870 with a final mass concentration of 0.05%, trehalose with a ...

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Abstract

The invention provides a colloidal gold labeling method for improving sensitivity as well as application and a product thereof, and relates to the technical field of immunochromatography. The colloidal gold labeling method provided by the invention comprises the following steps: uniformly mixing to-be-labeled protein treated by a protein activator with a colloidal gold solution, and carrying out labeling reaction to obtain gold-labeled protein. Wherein the protein activating agent comprises a nonionic surfactant, a saccharide protective agent and a reducing agent, and through the synergistic cooperation of the components, the interaction between the gold-labeled conjugates is effectively reduced, aggregation is avoided, the stability of the gold-labeled conjugates is improved, and the protein activity is protected; the pH value of the colloidal gold solution is 0.3-0.8 lower than the isoelectric point of the to-be-labeled protein, the protein is labeled under the condition, and the formed gold-labeled conjugate is good in stability and high in labeling efficiency. The method is easy and convenient to operate, the prepared gold-labeled conjugate is high in labeling efficiency and good in stability, and the prepared kit is high in sensitivity and not prone to missing detection.

Description

technical field [0001] The invention relates to the technical field of immunochromatography, in particular to a colloidal gold labeling method for improving sensitivity and its application and product. Background technique [0002] The combination of colloidal gold and protein mainly depends on three forces: electrostatic force, hydrophobic force and gold-sulfur bond. The main force of colloidal gold binding to protein is different at different pH, which affects the labeling efficiency and the stability of the conjugate. [0003] When the pH value is lower than the isoelectric point of the protein, the electrostatic interaction is the main force, the protein is positively charged, and the colloidal gold is negatively charged. The phenomenon of gold, the labeling efficiency is high but the stability of the conjugate formed is poor. [0004] When the pH is higher than the isoelectric point of the protein, the protein is negatively charged, and the negative charge of the gold...

Claims

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Application Information

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IPC IPC(8): G01N33/543G01N33/68G01N33/558G01N33/52G01N33/531G01N33/569
CPCG01N33/54313G01N33/68G01N33/558G01N33/52G01N33/531G01N33/56983G01N2333/005
Inventor 魏鹏海朱兆奎储寒君梁权张淑芳
Owner SHANGHAI BIOGERM MEDICAL TECH CO LTD
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