Protein markers of gestational diabetes mellitus in urine and use in early diagnosis
A gestational diabetes and protein technology, which is applied in disease diagnosis, mass spectrometry, biological testing, etc., can solve the problems of sensitivity and specificity to be considered, unrecognized, and poor specificity, and achieve the goal of overcoming antibody specificity and titer Limited, high selectivity, good reproducibility
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Embodiment 1
[0091] Example 1: Detection of gestational diabetes-related proteins in urine
[0092] We screened urine for gestational diabetes-associated proteins using Data independent acquisition (DIA).
[0093] Materials and Reagents
[0094] 1) Instrument: Orbitrap Fusion Lumos Tribrid mass spectrometer (Thermo Scientific Company).
[0095]2) Main reagents: trypsin (Promega Company); C18 solid-phase extraction cartridge (3CC, 60 mg, Waters Company); C18 reverse-phase chromatographic column (4.6 mm×250 mm, C18, 3 μm, Waters Company).
[0096] 3) Samples: urine from 15 patients with gestational diabetes (6-10 weeks of pregnancy) and 15 normal controls from Peking Union Medical College Hospital.
[0097] 1.1 Collection of human urine samples and enrichment of urine protein
[0098] Collect fasting morning urine and centrifuge at 5000g for 30min to remove sediment. The supernatant was precipitated with ethanol overnight at 4 to precipitate the protein. Reconstitute the protein pellet ...
Embodiment 2
[0105] Example 2: Mass spectrometric detection of urine protein NOV protein homologues, peroxide reductase-5, and hepatocyte growth factor-like proteins.
[0106] In order to better detect the application of these differential proteomic analysis results in further larger clinical samples, we used targeted mass spectrometry to monitor the differential proteins identified. Targeted analysis (parallel reaction) monitoring is a target mass spectrometry quantitative analysis technology based on the secondary mass spectrometry signal. Compared with the traditional selective reaction monitoring technology, it does not need to pre-design the parent ion / product ion pairing information of the targeted protein, saving Experimental design and operating time; and higher selectivity, better sensitivity, better reproducibility, and stronger anti-interference ability in complex backgrounds. Compared with immunization methods, it is no longer subject to commercial antibodies, and overcomes the...
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