New method and product for detecting tuberculosis antibody in serum sample

A tuberculosis antibody and antibody detection technology, applied in measuring devices, instruments, fluorescence/phosphorescence, etc., can solve the problems of lack of models and evaluations, and achieve good consistency results

Inactive Publication Date: 2009-09-16
CHINESE ACAD OF INSPECTION & QUARANTINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, whether the suspension chip can detect tuberculosis antibodies in human serum, and its quantitative detection ability, there is still a lack of models and evaluations.

Method used

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  • New method and product for detecting tuberculosis antibody in serum sample
  • New method and product for detecting tuberculosis antibody in serum sample
  • New method and product for detecting tuberculosis antibody in serum sample

Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0045] 2. Preparation of samples to be tested

[0046] 1. Preparation of target analyte samples

[0047] The target analyte is rabbit anti-tuberculosis IgG, and the interfering sample or the sample used as method-specific test is other antibodies or other proteins other than the target detection object, including mouse anti-influenza NP IgG, rabbit anti-SARS IgG, rabbit anti-avian influenza H5N1 serum, Rabbit anti-plague IgG, BSA, casein, tryptone, etc. All the above-mentioned samples to be analyzed were dissolved in the sample diluent and stored at -4°C. The stock solution concentration of rabbit anti-tuberculosis IgG is 0.009mg / mL. In the comparison experiment, the same sample was used for the detection of ELISA and suspension chip.

[0048] Dilute the rabbit anti-tuberculosis IgG sample diluent to be analyzed into samples of different concentrations in a 4-fold ratio to draw a standard curve for sample detection dose-response. Among them, the concentration of several sam...

Embodiment 1

[0051] Embodiment 1, the preparation of the protein suspension chip that detects tuberculosis antibody

[0052] 1. Capturing antigen-coated encoded microspheres

[0053] The No. 027 coded microspheres used in the present invention were purchased from BIO-RAD Company of the United States. The coded microspheres are used to label the tuberculosis 16KD protein and 38KD protein antigens that can capture avian influenza antibodies, that is, the tuberculosis 16KD protein and 38KD protein are used to coat the microspheres.

[0054] A. Activation of encoded microspheres

[0055] Take 100μL (1.25×10 6 pcs) encoded microspheres into a 1.5mL centrifuge tube, centrifuge at 14000×g, carefully aspirate and discard the supernatant. Add 100 μL of microsphere washing buffer to suspend, shake and sonicate, centrifuge at 14000×g, carefully aspirate and discard the supernatant. Add 100 μL of microsphere activation buffer, then add 10 μL of freshly prepared EDC (50 mg / mL), then add 10 μL of fre...

Embodiment 2

[0065] Embodiment 2, optimization of suspension chip preparation method conditions

[0066] 1. Selection of the amount of antigen coating on microspheres

[0067] 100 μL of microspheres coded as No. 027 were coated with 1 μg, 3 μg, 6 μg, 9 μg, 12 μg, 15 μg, and 20 μg, respectively. After testing the effect comparison, with 6μg / 1.25×10 6 A microsphere, that is, 12-24ng / 2500-5000 microspheres / test coating, has the best coating effect. After counting under a microscope, store it in a dark place and refrigerate it for later use. Such as figure 1 As shown, the No. 027 microspheres coated with tuberculosis 16KD protein and 38KD protein antigens all fell in the correct detection area, and obtained high signal-to-noise ratio results (MFI value is much greater than 2000), indicating that the optimized suspension chip detection system can be successful For the detection of tuberculosis antibodies.

[0068] 2. Optimization of biotinylated antibodies

[0069] The present invention re...

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Abstract

The invention relates to a new quantitative detection method and a product for detecting tuberculosis antibody in a serum sample. The method of the invention has good detection capability, high sensitivity, strong specificity and wide dynamic range. The invention establishes an open detection modularised platform of pathogenic bacteria antibody protein suspension chip detection with the tuberculosis antibody as a representative.

Description

technical field [0001] The invention relates to a novel quantitative detection method and product for detecting tuberculosis antibodies in serum samples and a preparation method thereof. Background technique [0002] Mycobacterium tuberculosis (M.tuberculosis, TB), commonly known as Mycobacterium tuberculosis, is the pathogenic bacteria that causes tuberculosis. It can invade all organs of the body, but tuberculosis is the most common. Tuberculosis is still an important infectious disease. An estimated one-third of the world's population is infected with Mycobacterium tuberculosis. According to the WHO, about 8 million new cases occur each year and at least 3 million people die from the disease. Before the founding of the People's Republic of my country, the mortality rate reached 200-300 people / 100,000, ranking first among the causes of death from various diseases. Morbidity and mortality are greatly reduced. However, it should be noted that in some areas of the world, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/569G01N33/543G01N21/64G01N33/545
Inventor 王静杨永莉孙肖红杨宇胡孔新
Owner CHINESE ACAD OF INSPECTION & QUARANTINE
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