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Blood group antigen chip and application thereof in detection of unexpected erythrocyte antibodies

A technology of erythrocytes and erythrocyte membranes, which is applied in the field of biomedicine, can solve the problems of poor sensitivity of erythrocyte accidental antibodies, low throughput screening ability, and narrow dynamic linear range, so as to avoid complex operation steps, wide linear range, and high sensitivity. Effect

Active Publication Date: 2018-01-30
GENERAL HOSPITAL OF PLA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in the above-mentioned patents, there are generally technical problems such as poor sensitivity, narrow dynamic linear range, low throughput screening ability, serious false negative problems, and inconvenient storage of detection reagents in the detection of unexpected antibodies in red blood cells in clinical blood.
[0006] Patent CN200880019301.1 realizes in vitro identification of individual anti-erythrocyte antibodies by immobilizing red blood cells or red blood cell membrane fragments on beads. Changes in the coating method, the concentration of the secondary antibody, and the dilution concentration of the sample will all affect the accuracy of the detection, so the method needs to be redesigned and evaluated

Method used

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  • Blood group antigen chip and application thereof in detection of unexpected erythrocyte antibodies
  • Blood group antigen chip and application thereof in detection of unexpected erythrocyte antibodies
  • Blood group antigen chip and application thereof in detection of unexpected erythrocyte antibodies

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] Example 1 Effect of Microsphere Coupling Conditions on Detection of Unexpected Antibodies in Erythrocytes

[0060] Select three kinds of uncoupled microspheres and resuspend them by ultrasonication for 2 minutes, take 50 μl concentration of 2.5×10 6microspheres / ml into the EP tube, place the EP tube on the magnetic plate for 60 seconds, and remove the supernatant; remove the magnetic separator and resuspend the microspheres with 100 microliters of deionized water, and place the EP tube on the magnetic plate Plate for 60 seconds, remove the supernatant; remove the magnetic separator and use 80 microliters of 100mM, pH6.2 disodium hydrogen phosphate buffer, shake to resuspend the microspheres, add 10 microliters of 50mg / ml Sulfo-NHS solution (dissolved in water), gently shake and mix, add 10 microliters of 50mg / ml EDC solution (dissolved in water), gently shake and mix, incubate at room temperature for 20 minutes, gently shake and mix once every 10 minutes; Place the EP ...

Embodiment 2

[0062] Example 2 The influence of the dilution factor of the detection sample on the detection of unexpected antibodies in red blood cells

[0063] Get 10 examples of positive specimens (detected by the microcolumn gel method) and 10 examples of negative specimens (detected by the microcolumn gel method), and carry out 10-fold, 30-fold and 90-fold dilutions of the 20 specimens respectively and prepare them in Example 1 The coupled microspheres were reacted, and the specific steps were: take a 96-well plate, add 200 microliters of PBS-TBN solution to each well and incubate at 37°C for 1 hour to block. Add 50 microliters of antibodies with different dilution factors and 2500 coupled microspheres, make up the volume of the system to 100 microliters for reaction; centrifuge the microplate at 2000rpm for 2 minutes, then place the 96-well plate on the magnetic plate for 2 minutes, Remove the supernatant; wash the 96-well plate twice with 100 microliters of PBS-TBN solution; add 50 m...

Embodiment 3 2

[0065] Example 3 The impact of the added concentration of the secondary antibody on the detection of unexpected antibodies in erythrocytes

[0066] Get 10 cases each of positive samples (detected by microcolumn gel method) and negative samples (detected by microcolumn gel method), and react with the coupled microspheres prepared in Example 1, wherein 8ug / ml of Cy3-labeled anti-human IgG and 2ug / ml Cy3-labeled anti-human IgG were used as secondary antibodies. The specific steps included: taking a 96-well plate, adding 200 microliters of PBS-TBN solution to each well and incubating at 37°C for 1 hour to block. Add 50 microliters of antibody with the same dilution factor and 2500 coupled microspheres, make up the volume of the system to 100 microliters for reaction; centrifuge the microplate at 2000rpm for 2 minutes, then place the 96-well plate on the magnetic plate for 2 minutes, Remove the supernatant; wash the 96-well plate twice with 100 microliters of PBS-TBN solution; add ...

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Abstract

The invention discloses a blood group antigen chip and application thereof in detection of unexpected erythrocyte antibodies. The detection comprises: (1) coating a carrier with a blood group erythrocyte membrane antigen; (2) contacting a tested sample with the antigen of the carrier; (3) adding a second antibody labeled with a marker; and (4) performing signal detection on the marker. The blood group antigen chip and a detection method of unexpected erythrocyte antibodies are high in sensitivity, good in accuracy, independent of erythrocytes, and capable of detecting binding antibodies of a plurality of rare blood group antigens.

Description

technical field [0001] The invention relates to the technical field of biomedicine, in particular to the serological detection technology of blood group antibodies, in particular to a blood group antigen chip, its preparation method and its application in the detection of red blood cell accidental antibodies. Background technique [0002] Blood transfusion safety has become an important issue in medical and health work, and has attracted great attention from the whole society. The World Health Organization (WHO) has always attached importance to blood transfusion safety work, and has further strengthened blood safety work in recent years. Blood safety has been listed by WHO as one of the seven key tasks of global health work. Blood transfusion safety issues include the issues of infectious adverse transfusion reactions and non-infectious adverse transfusion reactions. The United Kingdom, the United States, and France have all launched the blood transfusion serious harm repo...

Claims

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Application Information

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IPC IPC(8): G01N33/80G01N33/543
CPCG01N33/543G01N33/546G01N33/551G01N33/80
Inventor 汪德清杨璐于晓波
Owner GENERAL HOSPITAL OF PLA
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