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Four causative agent differential immunoglobulin G antibody and affinity index testing kit

A technology of immunoglobulin and detection kits, which is applied in the field of medical and biological detection, and can solve the problems of single detection of IgM antibody and unreasonableness

Inactive Publication Date: 2006-07-26
国家人口和计划生育委员会出生缺陷干预工程技术中心 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] (2) Distinguish the test objects: At present, in many areas of China, non-pregnant women of childbearing age and pregnant women are all tested for IgM antibodies related to these pathogens. It is obviously unreasonable to do so.
If the relevant IgG antibodies have not been detected before pregnancy, it is easy to distinguish according to the results of the relevant IgM detection after pregnancy. Do the IgM test, but the IgM positive test alone cannot distinguish between primary and secondary reinfection, so it is necessary to test the affinity index of IgG for IgM positive patients

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Embodiment 1: Rubella virus-specific IgG antibody detection kit and its preparation

[0042] (1) Antigen preparation

[0043] 1. Cultivate Vero cells to a monolayer, and inoculate rubella virus Gos strain, the cell growth medium is DMEM culture medium containing 10% calf serum, after 1-2 hours of adsorption at 37°C, the inoculum is poured out, and the inoculum is added with 1% small calf serum DMEM maintenance solution with bovine serum, 35°C, 5% CO 2 Culture for 5-7 days.

[0044] 2. Collect the infected cells, centrifuge to get the supernatant and add NaCl and PEG (MW=6000).

[0045] 3. Centrifuge to get the precipitate.

[0046] 4. Purified by sucrose gradient centrifugation at 4°C, 85000g, 2h.

[0047] 5. Take the middle zone and test the protein after dialysis, determine the coating concentration by titration of yin and yang serum, subpackage, and store at -70°C.

[0048] (2) ELISA coated solid phase carrier

[0049] 1. Dilute the Rubella antigen to the optim...

Embodiment 2

[0059] Example 2: Rubella virus-specific IgG antibody affinity index detection kit and its preparation

[0060] (1) Antigen preparation: same as Example 1

[0061] (2) ELISA coated solid phase carrier: same as Example 1

[0062] (3) Operation process

[0063] 1. Detection of serum: 1:101 dilution, 100 μl per well, two wells for each sample, 37 ° C, 60 min.

[0064] 2. After washing twice with the lotion, add 300 μl of denaturant to one well of each sample and keep it for 10 minutes, add 300 μl of lotion to the other well and keep it for 10 minutes, then shake it off, and wash twice with the lotion.

[0065] 3. Add enzyme-labeled anti-human IgG, 100 μl per well, 37°C, 60min.

[0066] 4. After washing five times, add TMB-H 2 o 2 Chromogenic system, room temperature, dark, 30min.

[0067] 5. Add stop solution (2M H 2 SO 4 ), measure the OD value of each well with a microplate reader at 450 nm.

[0068] 6. Calculate the affinity index of each specimen according to the for...

Embodiment 3

[0071] Example 3: CMV-specific IgG antibody detection kit and its preparation

[0072] (1) Antigen preparation

[0073] 1. Human embryonic diploid cells were cultured to a monolayer and inoculated with CMV-AD 169 strain, the cell growth medium is DMEM culture medium containing 10% calf serum, after 1-1.5 hours of adsorption at 37°C, the inoculum is poured out, and DMEM maintenance solution containing 1% calf serum is added, at 35°C, 5% CO 2 Grow 5-6 days, 100% lesion, trypsinize.

[0074] 2. Collect the infected cells, add 0.05M / L glycine-NaOH, pH9.0 buffer to suspend.

[0075] 3. After ultrasonic treatment, centrifuge to get the supernatant.

[0076] 4. Measure the protein, determine the coating concentration by yin and yang serum checkerboard titration, and store in -70°C.

[0077] (2) ELISA coated solid phase carrier: same as Example 1

[0078] (3) operation process: with embodiment 1

[0079] (4) kit composition: with embodiment 1

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Abstract

A reagent kit of specificity IgG and its affine index detection is prepared by planting giant cell virus AD169 plant, herpes simplex virus I type SM-44 plant, herpes simplex II type G pland on diploid cell of human embryo lungs, planting nettle rash virus Gos plant, bowworm RH plant on Vero cell, obtaining antigen Via developing, encapsulating polystrene ELISA reaction plate after tiration and purification to be certain concentration as well as preparing specimen dilution liquid developer solution, concentration cleaning liquid and so on.

Description

technical field [0001] The invention belongs to the field of medical and biological detection. Specifically, the invention relates to four pathogen-specific IgG antibodies of giant cells, rubella, herpes simplex and toxoplasma gondii and an affinity index detection kit and a preparation method thereof. Background technique [0002] Preventing the birth of children with birth defects is of special significance to China. Intrauterine infection is one of the important reasons for the birth of children with birth defects. In recent years, the family planning systems in various regions of China have gradually carried out work on the prevention of intrauterine infection. The detection of some diseases that may cause intrauterine infection and birth defects is currently mainly focused on the detection of cytomegalovirus (CMV), rubella virus (RV), herpes simplex virus (HSV) and toxoplasma gondii (Toxo) infection. Most of the methods are to detect the corresponding specific antibodi...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/53G01N33/569
Inventor 马旭侯林浦姜英涛高英
Owner 国家人口和计划生育委员会出生缺陷干预工程技术中心
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