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Quantitative detection kit for Sflt-1

A quantitative detection and kit technology, applied in the field of kits, can solve the problem of low sensitivity and achieve the effect of high sensitivity, good repeatability, and small sample volume

Inactive Publication Date: 2020-05-12
NINGBO AUCHEER BIOTECHNOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, the detection methods of sFLT-1 include enzyme-linked immunoassay and colloidal gold colorimetric method. low sensitivity

Method used

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  • Quantitative detection kit for Sflt-1
  • Quantitative detection kit for Sflt-1
  • Quantitative detection kit for Sflt-1

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] A quantitative detection kit for Sflt-1, comprising a substrate 1 and a cover plate 2, the substrate 1 and the cover plate 2 are formed by bonding, the cross sections of the substrate 1 and the cover plate 2 are rectangular structures, and the cover plate 1 is provided with a sampling hole 3 and a detection hole 4, and the substrate 1 is sequentially provided with a sampling area 5, a reaction area 6, a detection area 7, and a waste liquid area 8, and the reaction area 6 is coated with fluorescent microspheres Labeled sFLT-1 antibody I, sFLT-1 antibody II coated with different epitopes is immobilized in the detection area 7, the lower end of the sample application area 5 is provided with an opening connected to the upper end of the filter channel 9, and the reaction area 6 The upper end is provided with an opening to communicate with the lower end of the filter channel 9, the filter channel 9 is provided with multi-layer filter cotton 10, the lower end of the reaction zo...

Embodiment 2

[0028] In this example, the preparation method of the microfluidic chip for quantitative detection of sFLT-1 is described in detail, which specifically includes the following steps:

[0029] (1) fixing the multi-layer filter membrane in the reaction channel;

[0030] (2) Wash the fluorescent microspheres with 0.05mol / L MES buffer (pH=7.2). After washing, add carbodiimide and N-hydroxysuccinimide, and react at room temperature for 2 hours to obtain a fluorescent microsphere solution. The sFLT-1 antibody I was dissolved in 0.05mol / L PBS buffer (pH=7.2) to obtain the sFLT-1 antibody I solution, and the fluorescent microsphere solution was added to the sFLT-1 antibody I solution, reacted at room temperature for 2 hours, and centrifuged. Wash 3-5 times with 0.01mol / L PBS buffer (pH=7.2) to obtain sFLT-1 antibody I labeled with fluorescent microspheres, and apply the sFLT-1 antibody I labeled with fluorescent microspheres dropwise on the reaction area;

[0031] (3) Immobilize and c...

Embodiment 3

[0035]Performance analysis of this kit:

[0036] 1. Linearity: The kit uses the concentration value of the sFlt-1 standard substance (0, 10, 50, 400, 2000, and 10000 pg / mL for the standard substance) as the X-axis, and the signal value of the standard substance as the Y-axis to establish a calibration Curve, according to the intensity of the signal value of the sample to be tested back to calculate the corresponding concentration value, such as Figure 4 As shown, the linear equation is y=246.44x-9948.5, R 2 =0.996, the lowest detection limit is 10pg / mL.

[0037] Get the sFlt-1 standard substance that concentration is 10pg / ml and carry out 10 tests, utilize linear equation y=246.44x-9948.5, calculate detection concentration value; Detection result is as shown in table 1, and the average value of determination is respectively 9.994pg / ml , the standard deviation was 0.3997, and the intraassay coefficient of variation was 4%.

[0038] Table 1

[0039] sFlt-1 stan...

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Abstract

The invention discloses a quantitative detection kit for Sflt-1. The kit comprises a substrate and a cover plate, the substrate and the cover plate are formed by bonding, a sample adding hole and a detection hole are formed in the cover plate, a sample adding area, a reaction area, a detection area and a waste liquid area are sequentially arranged on the substrate, the reaction area is coated witha fluorescent microsphere labeled sFLT-1 antibody I, and the detection area is internally and fixedly coated with sFLT-1 antibodies II with different epitopes. The kit is high in sensitivity, good inrepeatability, extremely small in required sample size, short in detection time and simple and convenient to operate, the operation process can be greatly simplified, the consumption of samples and reagents is reduced, expensive instruments do not need to be arranged, on-site instant detection becomes possible, and the kit has huge economic value and social value; meanwhile, a doctor can be helped to diagnose related diseases of preeclampsia, and early identification and intervention are performed on high-risk groups, so that the safety of maternal and infant in the gestation period is guaranteed.

Description

technical field [0001] The invention relates to the technical field of kits, in particular to a quantitative detection kit for Sflt-1. Background technique [0002] The ectodomain of Flt-1 is spliced ​​to form soluble vascular endothelial growth factor receptor-1 (sFlt-1), soluble vascular endothelial growth factor receptor 1 (sVEGFR-1), also known as soluble fms-like tyrosine kinase-1 (sFLT -1), originally discovered in human umbilical vein endothelial cells, is a glycoprotein with tyrosine kinase activity. sFlt-1 retains the high-affinity properties combined with PLGF and VEGF, and can make the tyrosine of PLGF and VEGF Acid kinase activity is lost, so it is believed that sFlt-1 is an antagonist of PLGF and VEGF. sFlt-1 is associated with the onset of preeclampsia. sFlt-1 plays an important role in the pathogenesis of preeclampsia, and its high expression can reduce the expression levels of PLGF and VEGF, leading to angiogenesis disorders and endothelial cell damage, whi...

Claims

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Application Information

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IPC IPC(8): G01N33/74G01N33/573G01N33/543
CPCG01N33/54313G01N33/573G01N33/74G01N2333/71G01N2333/9121G01N2800/368
Inventor 周义正黄丹娣陈星星董文霞刘超波崔丛丛
Owner NINGBO AUCHEER BIOTECHNOLOGY CO LTD
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