Immunochromatographic reagent strip for fluorescent quantitative detection of PAPP-A (Pregnancy Associated Plasma Protein A) and preparation method of reagent strip

A fluorescent quantitative detection, PAPP-A technology, applied in the field of immunodiagnosis, can solve the problems of many influencing factors, high price, long time consumption, etc., to achieve the effect of ensuring accuracy, simple operation and high sensitivity

Inactive Publication Date: 2017-11-07
SHENZHEN YHLO BIOTECH +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

ELISA method has many detection steps, takes a long time, and there are many influencing factors in the operation process, which can easily cause false positive and false negative results
Therefore, it is gradually replaced by the chemiluminescence method, but this method is a fully closed system, which is expensive and requires special training for instrument users. The maintenance and testing costs are high, and it is not suitable for single-person and small-batch testing. It is not conducive to PAPP at present. -A detection is

Method used

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  • Immunochromatographic reagent strip for fluorescent quantitative detection of PAPP-A (Pregnancy Associated Plasma Protein A) and preparation method of reagent strip

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Preparation of PAPP-A fluorescent immunochromatographic test strips:

[0033] (1) Preparation of fluorescent microsphere-labeled protein

[0034]Take 0.1 mL of 10% fluorescent microspheres, centrifuge at 15,000 rpm for 15 minutes, adjust the concentration of the precipitate to 1% with 50 mM pH 6.5 MES buffer, and ultrasonically disperse; add carbodiimide (EDC) at a final concentration of 2 mg / mL ), mix well, then add N-hydroxysuccinimide (NHS) with a final concentration of 5 mg / mL, mix well; incubate at room temperature for 20 minutes and then centrifuge at 15,000 rpm for 15 minutes, and the precipitate is washed with 50 mM pH6.5 MES buffer dissolve. Ultrasonic disperse the reconstituted fluorescent microspheres, add 0.2 mg PAPP-A monoclonal antibody and 0.1 mg avidin respectively in two tubes, mix well, rotate and mix at room temperature for 2 hours, centrifuge at 15,000 rpm for 15 minutes, and use the precipitate Reconstitute in Tris-HCl (20 mM, pH 8.0) containing 3...

Embodiment 2

[0043] Quantitative detection of pregnancy-associated protein A (PAPP-A) concentration in blood samples by fluorescence immunochromatography

[0044] (1) Standard curve drawing

[0045] Prepare PAPP-A antigen with negative plasma to 1000 ng / mL, 500 ng / mL, 100 ng / mL, 50 ng / mL, 30 ng / mL, 10 ng / mL, 0 ng / mL, use the same batch of reagents, Each concentration point was tested 6 times. Take the fluorescence intensity ratio of the detection line (T band) and the quality control line (C band) as the ordinate, and the concentration of PAPP-A reference substance as the abscissa, establish an equation and fit it into a standard curve, and use the programming software to record the standard curve information Write to the ID chip.

[0046] (2) Detection of samples:

[0047] Take out the test strip from the kit, tear open the aluminum foil bag, lay the test strip flat, and equilibrate for 5 minutes. Take 100 μL of sample and add it to the sample well, and react at room temperature for 15...

Embodiment 3

[0050] Please refer to figure 1 , an immunochromatographic test paper for fluorescent quantitative detection of PAPP-A, the immunochromatographic test strip includes a PVC base plate 7, and the PCV base plate 7 is sequentially provided with a sample pad 1, a marking pad 2, and a coating film 3. Absorbent paper 4, the marking pad 2 is connected to the sample pad 1, the coating film 3 is connected to the marking pad 2, the absorbent paper 4 is connected to the coating film 3 are connected; the PAPP-A monoclonal antibody labeled with fluorescent microspheres and avidin labeled with fluorescent microspheres are sprayed on the marker pad 2; detection lines 5 and quality control lines 6 are provided on the coating film 3, The detection line 5 and the quality control line 6 are separated by 4-8 mm, and the detection line 5 is coated with another PAPP-A monoclonal antibody that is at a different epitope from the PAPP-A monoclonal antibody labeled with the fluorescent microspheres , t...

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Abstract

The invention discloses an immunochromatographic test strip for fluorescent quantitative detection of PAPP-A (Pregnancy Associated Plasma Protein A) and a preparation method of the test strip, and belongs to the technical field of immunologic diagnosis. In order to overcome the defects in the prior art, the immunochromatographic test strip for the fluorescent quantitative detection of the PAPP-A adopts the following technical scheme that the test strip comprises a sample pad, a marker pad, a coating film and absorbent paper which are sequentially overlapped on a PVC (Polyvinyl Chloride) bottom plate; avidin is sprayed on the marker pad; a quality control line is coated with a rabbit anti avidin antibody for specific recognition of the avidin. The test strip has the following advantages that the test strip is used for evaluating and monitoring a level of the in-vivo PAPP-A, is simple to operate, quick, convenient, economic and accurately quantitative, and is more suitable for wide development in various medical institutions.

Description

technical field [0001] The invention belongs to the technical field of immunodiagnosis, and in particular relates to an immunochromatographic test strip for fluorescent quantitative detection of pregnancy-associated protein A (PAPP-A) and a preparation method thereof. Background technique [0002] Pregnancy associated plasma protein A (PAPP-A) is a macromolecular glycoprotein derived from the placenta, which is synthesized by placental syncytiotrophoblast cells, decidual cells and endometrial stromal cells in normal menstrual cycle. It is secreted into the serum, and its concentration increases with the increase of gestational age until term. It has been proved that the concentration of PAPP-A in the serum of pregnant women with Down syndrome (DS) fetuses in the first trimester is significantly reduced, so PAPP-A is widely recognized as an effective indicator for DS screening, and its concentration reduction indicates that the newborn has Down syndrome. increased risk of Sy...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/558G01N33/577G01N33/533G01N21/64
CPCG01N33/689G01N21/6402G01N21/6428G01N33/533G01N33/558G01N33/577G01N2021/6439G01N2800/368
Inventor 钱纯亘邹畅夏福臻张赛胡鹍辉宋永波
Owner SHENZHEN YHLO BIOTECH
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