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Detection method for rapidly and quantitatively detecting content of antinuclear antibody

An anti-nuclear antibody and detection method technology, applied in the field of rapid quantitative detection of anti-nuclear antibody content, can solve the problems of inaccurate quantification, false positive target antigen, membrane strip pollution, etc., to help monitor the disease and curative effect, predict the occurrence of and prognosis, the effect of accurate quantitative detection

Pending Publication Date: 2019-02-12
QINGDAO UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

The more antigens coated on the detection membrane strip, the more times it is pasted, the greater the possibility of contamination of the membrane strip, which makes the test results of the detection membrane strip poor in repeatability, and may also cause misreading of the test results due to pasting errors
Although this blotting method has the advantages of strong specificity, easy operation, and relatively objective results, it can only make qualitative (negative, positive, strong positive) judgments on the serum antibody levels of patients, and cannot be accurately quantified, so it cannot be used. Reflect the outcome of the disease and make a prognosis judgment
In addition, in clinical practice, this test item has a certain false positive and false negative rate for the detection of some target antigens, and the cost is high, so its reference value and application for clinical diagnosis have certain limitations

Method used

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  • Detection method for rapidly and quantitatively detecting content of antinuclear antibody
  • Detection method for rapidly and quantitatively detecting content of antinuclear antibody
  • Detection method for rapidly and quantitatively detecting content of antinuclear antibody

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] Step 1. Sample layout of Wes test board

[0055] The components of the detection reaction are pre-added to the detection plate of the Wes system, and the detection reaction is carried out in the matching cartridge containing 25 capillaries.

[0056] The layout of the samples in the test plate (25 wells / row, 1-25 wells from left to right) is as follows:

[0057] The first line, line A: A1 is the molecular weight standard, A2-A25 is the multi-combination target antigen sample;

[0058] The second line, line B: all are blocking solutions;

[0059] The third row C: C1 is the blocking solution, C2-C25 are the diluted serum samples;

[0060] The fourth row D: D1 is HRP-labeled streptavidin-HRP, D2-D25 is the secondary antibody-HRP mixture;

[0061] The fifth line, line E: all are luminescent substrate chromogenic solutions;

[0062] The sixth line, line F: empty.

[0063] Blocking solution, Streptavidin-HRP, and luminescent substrate chromogenic solution were equipped wi...

Embodiment 2

[0081] Step 1. Normalization processing of quantitative data

[0082] The peak area value (peakarea value, Pav) of the optical signal product peak area value (peakarea value, Pav) of the internal standard polypeptide control antigen sys ctrl in each capillary was 65,000, which was used as the control internal reference for normalizing the data of each antinuclear antibody (ANA). The formula for calculating the peak value of the ANA normalized product in serum samples is as follows:

[0083]

[0084] The normalized Pav value can be used as a quantitative standard unit for the antibody content level in serum.

[0085] Step 2. Establish the sample dilution factor

[0086]As the concentration of antinuclear antibody increases, the signal intensity of the luminescent substrate increases, but limited by the detection sensitivity and detection dynamic range of the detection system for the brightness of the band, the detection results of samples with too low or too high levels are...

Embodiment 3

[0098] Step 1. Optimal screening positive cut-off value (Cut-off value)

[0099] Receiver operator characteristic curve (ROC curve) is mainly used to select the best signal detection model or set the best threshold in the same model. In this embodiment, the ROC curve is used to set the best threshold.

[0100] Adopt the operating steps of Example 1, with the normalization processing method of Example 2, the sample dilution factor is taken 1:100, obtain the signal-to-noise ratio of six antinuclear antibodies in the patient sample (positive) and the control sample (negative) respectively S / N value and normalized Pav value.

[0101] Use SPSS data processing software to draw the ROC curve. The specific method is to use the Pav value as the data column, the positive and negative samples as the status column, and use "positive" as the positive status value to draw the ROC curve, and give many Cut- Sensitivity parameters and 1-specificity parameters corresponding to the off value, u...

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Abstract

The invention belongs to the technical field of biological detection analysis, and relates to a detection method for rapidly and quantitatively detecting content of an antinuclear antibody. A method which employs an ultramicro full-automatic protein expression quantitative analysis system and is simultaneously used for rapidly detecting content of each type of a plurality of antibodies in a detection sample is proposed. The method can have the advantages of rapid and accurate quantitative detection, high specificity, high sensitivity and the like, the content of the antinuclear antibody in patient serum can be quantitatively analyzed, and automation is more easily achieved. By the detection method, the quantitative grading standard for evaluating an antibody value of the sample and optimalscreening positive critical value are further built, and the detection method is used for forecasting or estimating whether a human body is probably developed with or already developed with various autoimmune diseases.

Description

technical field [0001] The invention belongs to the technical field of biological detection and analysis, and relates to a detection method for rapid and quantitative detection of antinuclear antibody content. Background technique [0002] Each autoimmune disease is accompanied by a characteristic autoantibody spectrum, which is an indispensable biological indicator for early diagnosis, disease assessment, disease staging, and prognosis judgment. Certain autoantibodies have become the gold standard for diagnosing corresponding diseases due to their high specificity and sensitivity for disease diagnosis. [0003] Antinuclear antibodies are a general term for autoantibodies that target the nuclear components of eukaryotic cells, and are a group of autoantibodies that are produced against DNA, RNA, protein, or molecular complexes of these substances in the nucleus. In clinical practice, among the mixed antigens used for auxiliary diagnostic detection, nRNP / Sm, Sm, SS-A, SS-B, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68
CPCG01N33/68
Inventor 王冰汤怀广王孔嘉丁怡心尹晓萌闫静宋晨璐
Owner QINGDAO UNIV
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