Biomarker for detecting nephritis and appliction thereof
A biomarker, nephritis technology, applied in the field of medicine, to achieve the effect of less dosage, less blood usage, and high sensitivity
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Embodiment 1
[0051] 1. The effect of different acids on serum microwave acid hydrolysis
[0052] (1) Randomly take 10 patient serums, take 2 of each serum, take 2μL to the microwave hydrolysis tube, add deionized water to 10μL, mix well and then add 10μL 6mol / L HCl or 4mol / L TFA;
[0053] (2) Under the conditions of power 100w and temperature 100℃, use a single-mode microwave proteolysis instrument for microwave acid hydrolysis for 10 minutes;
[0054] (3) Transfer the sample after microwave acid hydrolysis with 20 μL deionized water to a 1.5 mL centrifuge tube, repeat three times, and centrifuge to concentrate and remove the acid;
[0055] (4) Add 100μL methanol to each tube, centrifuge to concentrate to remove residual HCl, repeat three times;
[0056] (5) Each tube of the obtained dry sample is dissolved in 150μL deionized water at a speed of 13000r / min, centrifuged for 5min, and the supernatant is taken and used for anion exchange chromatographic analysis. After the analysis is complete, rinse t...
Embodiment 2
[0076] 1. Drawing the standard curve of monosaccharide
[0077] Accurately weigh the appropriate amount of fucose, galactosamine, glucosamine, galactose, glucose, mannose, add deionized water to prepare the above monosaccharides each containing 0.05mg / mL, 0.01mg / mL, 0.005mg / mL, 0.001 mg / mL, 0.0005mg / mL, 0.0001mg / mL, 0.00005mg / mL solution; take 80μL to the sample bottle for anion exchange chromatography analysis.
[0078] The chromatographic conditions are as follows:
[0079] Analytical column: Thermo Scientific Dionex Carbo PAC TM PA10, 4.0mm×250mm,
[0080] Guard column: Thermo Scientific Dionex Carbo PAC TM PA10, 4.0mm×50mm,
[0081] Eluent: 18mM NaOH for 0-18min; flow rate: 1.0mL / min, injection volume: 10μL, column temperature: 30℃,
[0082] Detector: electrochemical detector, gold electrode (P / N 061875), standard sugar potential, running time: 18min. Obtain the anion exchange chromatograms of 6 monosaccharide standards, see Image 6 A.
[0083] From Image 6 A can see that the 6 ...
Embodiment 3
[0096] Take 2μL of blood from 216 patients with diagnosed nephritis, and the rest of the steps are the same as in Example 2. Statistical test results, see Figure 7 .
[0097] will Figure 7 The results are summarized in Table 1, and the analysis results of the 6 monosaccharide concentrations are shown in Table 2.
[0098] Table 1. The relative trend of the 6 monosaccharides obtained after blood degradation in patients with nephritis and those without nephritis
[0099] Fucose Galactosamine Glucosamine Galactose glucose Mannose Nephritis ↑*** ↑*** ↑*** ↑** ↑** ↑***
[0100] Note: "NS" means no change compared with people without nephritis, "***p <0.001", "**p <0.01" and "*p "<0.05" respectively represent different significant differences compared with those without nephritis, and "↑" and "↓" respectively represent an increase or decrease compared with those without nephritis.
[0101] Table 2. Contrast results of 6 monosaccharide concentrations (μmol / L) in blood of patients with...
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