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Granular tag for biological detection

A biological detection and granular technology, applied in the direction of biological testing, material inspection products, etc., can solve the problems of cumbersome preparation process, multi-labeling, and long time-consuming, etc., and achieve the effect of high adsorption efficiency

Inactive Publication Date: 2005-09-28
朱有名
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The washing process takes more than ten hours, and most of the separation and passivation steps require chromatographic purification, which is very complicated.
It can be seen that the traditional method of preparing markers has four obvious disadvantages: 1. The preparation process is cumbersome and time-consuming; 2. The marker and the labeled substance are directly coupled through a coupling agent. The detection signal is relatively weak; 3. Due to the change of reaction conditions during the coupling process and the steric hindrance formed after labeling, it often has a certain impact on the original properties of the labeled components, such as the affinity of the labeled antigen or antibody , Reactivity decreased or disappeared, etc.; 4. Multiple labeling could not be performed

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0010] Example 1 Direct method

[0011] 1. Take 10-100ml of 0.01% chloroauric acid aqueous solution, add 1-5ml of 0.1-1% trisodium citrate, heat and boil for 5-50 minutes until the color becomes wine red, and the gold particle solution is obtained.

[0012] 2. Take 10ml of the above gold particle solution, and use 0.1mol / LK 2 CO 3 0.1mol / L HCL to adjust the pH to 6-11.

[0013] 3. Take a certain amount of HIV antigen and alkaline phosphatase solution, add it dropwise to the pH-adjusted solution, stir for 10-30 minutes, then add BSA to a final concentration of 0.1-1%.

[0014] 4. Centrifuge at 20000 rpm for 1 hour at low temperature, and discard the supernatant.

[0015] 5. Resuspend the pellet in a buffer solution containing preservatives and stabilizers, and store it at low temperature for later use.

Embodiment 2

[0016] Embodiment 2 indirect method

[0017] 1. Add 50-100ml of deionized water, 30-90ml of acetone, and 10-30g of styrene into a four-neck flask equipped with a stirrer, condenser tube, feeder and nitrogen gas guide device, and put it in a water bath at a certain temperature for 20- After 50 minutes, 0.2-0.5 g of potassium persulfate was added and reacted for 8-12 hours. The obtained emulsion was separated at a high speed of 25,000 rpm, and the precipitate was washed with buffer solution and then diluted to obtain latex particles.

[0018] 2. Add 5-10ml of 0.05-0.1mol / L n-aminocaproic acid and 10-20ml of 0.05-02mol / L pH7.2 buffer solution to 10ml of the above-mentioned latex particles.

[0019] 3. Take 2ml of the above solution, add 0.5-2ml of streptavidin (5000mg / L) and 5-15mg of carbodiimide, mix well, 37°C for 2 hours, add 0.1-0.5ml of buffer solution of PH8-10, After 30 minutes, centrifuge at 10,000 g for 1 hour, discard the supernatant, and wash the pellet twice with pH...

Embodiment 3

[0024] Example 3 Application of particulate markers in HIV antibody detection

[0025] 1. Take an ELISA plate pre-coated with HIV antigen, add 50ul of the serum to be tested to each well; add 50ul of positive control serum and negative control serum to the positive control well and negative control well respectively, and place at 37°C for 1h.

[0026] 2. After washing the plate 5 times with phosphate buffer containing surfactant, add 50 ul of the granular marker prepared according to Example 1 or Example 2 diluted in a certain proportion to each well, and place at 37° C. for 1 hour.

[0027] 3. After washing the plate 3 times with phosphate buffer solution containing surfactant, add 100ul of chromogen to each well and place at 37°C for 15min.

[0028] 4. After adding 50ul of stop solution to each well, place it on a microplate reader to measure the absorbance value of each well, and judge the result according to the absorbance value of each well.

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PUM

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Abstract

The present invention discloses one kind of granular marker for biological detection, and the granular marker is granular matter with at least one marking component and at least one marked component on the surface. The granular marker is prepared with granular matter as carrier and through adsorbing, combining or connecting the marking component and the marked component directly or indirectly. The present invention has simple granular marker preparing process and doubled detected signal of the marker, and makes it possible to connect several kinds of marking components and marked components onto the same kind of granular matter for multiple marking.

Description

technical field [0001] The invention relates to the technical field of biological detection. Background technique [0002] In the process of biological analysis and detection, in order to obtain detection signals or improve detection sensitivity, it is often necessary to combine a signal component or a component that is easy to generate a detection signal after a reaction, such as: luciferin, enzyme, biotin, isotope, etc., and another Components such as proteins, polypeptides, nucleic acids, etc. are coupled through chemical coupling agents, and the coupling products are traditional markers. If horseradish peroxidase (HRP) is used to label immunoglobulin IgG, the following five steps are generally required: 1. Treat HRP with fluorodinitrobenzene (FDNB) to block its α- and ε-amino groups to prevent HRP undergoes self-crosslinking; 2. Oxidize the carbohydrate groups of HRP with sodium periodate to make HRP a protein "aggregator" HRP-CHO with multiple aldehyde groups; 3. Let t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/48
Inventor 朱有名
Owner 朱有名
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