| [Objective]1. In patients with DVT filtered using two-dimensional electrophoresis of proteins in serum, exploratory search for deep vein thrombosis related differences in protein, using mass spectrometry for qualitative differences in protein。2. Using bioinformatics methods to analyze differences in protein function, find the difference of protein associated with thrombosis。3. Use Elisa technology to detect differences in protein in thrombosis associated with thrombus, the thrombus formation and expression of the change under normal condition. By analyzing qualitative quantitative differences in protein detection, the result of thrombosis. Hope for the next step development early prediction and diagnosis of deep vein thrombosis protein kit provides the necessary data and theoretical basis。[Materials and Methods]Part Ⅰ Patients in deep venous thrombosis screening of differential proteins and functional analysis。1.In the frist hospital attached to Kunming Medical University:;collecting of thrombosis in the outpatient and inpatient group of30patients with thrombosis (Group C) and dynamic observation of Orthopedic patients after major surgery, in the observed period (2weeks) did not include thrombosis of clots do not form a group (Group B) collected in30patients. From medical center to collect blood samples from volunteers in30cases of normal control group (group A)。2.From the three groups do dielectrophoresis,3patients were randomly each extraction of9pieces of electrophoresis film are compared; Using Image Master2D platinum5.0analysis software to analyze plastic figure, select the expression differences significant differences, its position on the gel. 3.Blood samples tested by the above solutions of proteins using matrix-assisted laser ionization-time of flight mass spectrometry (MALDI-TOF) technology, Combined with the informative Human Proteome database, identification of unknown proteins in the sample to determine its origin and nature。4. To identify the protein Pathway, pubmed protein database into bioinformatics analysis select protein related to DVT。Part II Validation has differences in screening protein (C4bBP) in patients with DVT expression changes in plasma。1.9cases in group A random sample; While in group B and group C use a random sample drawn10cases each。2. ELISA in detecting A, B, C group differences in protein expression; And analyze the changes of the differences in protein content in different groups.[Results]Part1Three groups of differental proteins filtered results and the analysis of function of differential proteins in the Group。1.9complete protein sample two-dimensional electrophoresis experiment, on9gel gel image analysis, select the10differences, which raised seven, cut the three.2. In thrombosis groups we chose four expression differences of protein mass spectrometry identification of points, respectively, Ig is A, the TpP, C4, C4bBP, found C4bBP associated with DVT. The function analysis is C4bBP combine with free type PS after leading to enhanced coagulation function。3. On thrombus formation in the group we screen out the point, there are differences between the four identified are respectively haptoglobin-like protein And Human Hemoglobi, haptoglobin, Chain A, Structure And Dynamics Of Human Apolipoprot ein C-Iii. Extract the two normal group differences, set by the mass spectrometry analysis of unnamed protein produc and beta-globin. For the identification of protein function analysis found that they have nothing to do with blood clots.Part2Validation expression changes of C4bBhP in plasma of patients with DVT.1,The results of ELISA C4bBP:Thrombosis group and normal control group and thrombosis has not formed the group; the concentration of C4bBP in plasma increased relative expression, The difference was statistically significant (P<0.05), Thrombosis is not shaped compared with normal control group, changes of C4bBP concentration in plasma was not obvious, the difference was not statistically significant (P>0.05)。[Conclusion]C4bBP in plasma of the patients with DVT expression and increased, its expression change trend and biological process with thrombosis of deep vein thrombosis, may be the early predictive diagnosis significance。... |
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