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Objective: To study the primary membranous nephropathy urine protein spectral characteristics, healthy urinary protein profiling to compare screened membranous nephropathy differentially expressed proteins, to explore mass spectrometry significance of the clinical diagnosis of the disease. : (N = 3) and idiopathic membranous nephropathy (n = 9) of patients in the morning urine specimens from healthy people, placed in -80 ℃ refrigerator for use in order to avoid bacterial growth and proteolysis within one hour. Dissolved completely, remove the 12 specimens with a diameter of 0.22 micron microporous ultrafiltration membrane filter to remove large impurities, and then -20 ° C pre-cooled acetone protein precipitation at -20 ° C refrigerator overnight at 4 ℃ 4000 × g centrifugation for 20 minutes, the supernatant was discarded, specimens of urine protein-containing precipitate was dried and concentrated in vacuo. Concentrated and dried protein precipitate was washed with PBS sufficiently dissolved, after the application of the Bradford method for the determination of protein concentration. Adjust the protein concentration, according to the measured specimen concentration take each case samples of 1 mg of protein mixed solid full enzymatic 1ug/ul, high performance liquid chromatography electrospray ionization tandem mass spectrometry (DETERMINATION OF TAXOL IN), the peptide The sequence results MASCOT software line database library that is used for the ipi.human, v3.05. Richard protein SEQ ID the Uniprot database of retrieval query name of the expressed protein, the role of the protein and cytological location and other relevant information. Results: healthy people 3 were identified 101 proteins, verified by hand, all the three specimens co-expression of 37 proteins. 9 urine samples of patients with membranous nephropathy were detected 174 proteins, all nine specimens co-expression of 20 proteins. Removal of normal human urine protein from a protein hand a total of 20 kinds of membranous nephropathy, but also the remaining 8 proteins, they are ceruloplasmin, lambda in chain C region, α-1 acid glycoprotein, vitamin D binding protein, leucine-rich alpha2 sugar protein receptors on ATP-sensitive inward rectifier potassium channel 11, the calcium-binding protein 39, IPI00745619 (unknown protein). Conclusion: with HPLC-ESI-MS/MS mass spectrometry method identified idiopathic membranous nephropathy urine protein, the polypeptide sequence spectrum of the disease, and ultimately get the 174 proteins. Ceruloplasmin, alpha2 sugar protein leucine-rich, sugar alpha1b protein, vitamin D-binding protein, calcium-binding protein 39 may be a biomarker of the disease.
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