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Objective To study the feasibility of detecting fetus RhD type gene by Surface Plasmon Reso-nance(SPR)technology,and to establish a new rapid diagnosis method for fetus RhD type gene.Methods The different types of DNA corresponding RNA probes were fixed on the surface of SPR chip by using amino cou-pling methods,and optimize the chip analysis condition,and then using the RNase H enzyme hydrolysis,signal amplification detection,at last the detection conditions were determined.We use the RhD type gene exon 5,7 of RNA probe to test its corresponding DNA molecules,and analyse the specificity and sensitivity of SPR chip detection signal.Results The SPR technique for detecting the exon 5,7 of RhD blood type gene shows good sensitivity and specificity in all,SPR technology can specifically detect the Exon 5,7 of RhD blood type gene, and the sensitivity of for detecting RhD gene exon 5,7 is 100 pmol/L by SPR.Conclusion The SPR technolo-gy can quickly detect RhD gene accordingly,SPR technology is simple,rapid,reliable and label-free,w hich can provide a new way predicting fetal RhD type for RhD negative prenatal.
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Objective To study the platelet antibody screening and crossing match by surface plasmon resonance(SPR) ,and to find a new way for platelet compatibility testing .Methods The corresponding universal platelet antigen was fixed on the SPR chip surface by the amino coupling method .Platelet antibody positive and negative control serum were analysed by SPR micro-ar-ray ,the stability ,sensitivity and specificity of this technique were discussed ,and compared with MAIPA assay .Finally we used the SPR technology to cross match for ten cases of the platelet antibody positive patients before infusion ,and to evaluate the effect of platelet infusion .Results For the SPR technology ,the stability ,sensitivity and specificity of platelet antibody detection were all better ,106 cases of the repeated platelet transfusion samples were tested by SPR assay and MAIPA method ,there was no signifi-cant difference between them(chi-square=0 .333 ,P>0 .05) ,the total consistency was 97 .2% .The 10 cases of platelet antibody positive patients were crossed match before platelet transfusion by SPR technology ,the good results of 8 cases of them were found by the clinical tracking evaluation ,1 h CCI>7 .5 ,24 h CCI>4 .5 .Conclusion SPR technology for screening platelet antibody mat-ches with MAIPA method in basic quality ,but SPR assay is simple ,rapid ,reliable and intuitive ,label free ,which can satisfy the re-quirements for clinical rapid detection of platelet antibody screening and crossing match .
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Objective To study the changes of the peripheral blood regulatory B cells and related cytokines in children patients with chronic idiopathic thrombocytopenic purpura (CITP),and to analyze their correlations with effective Th cells,and to discuss their roles in the pathogenesis of CITP.Methods The peripheral blood mononuclear cells (PBMC)were separated in 30 cases of CITP,then percentages of Th1 ,Th17,Th22 and Breg cells were detected by the flow cytometry(FCM).IFN-γ,IL-17,IL-22 and IL-10 levels in culture supernatant were measured by the ELISA method.Results Compared with the control group,the proportions of the peripheral blood Th1 ,Th17,Th22 subsets in CITP children were increased[(18.4±4.7)% vs.(10.82±4.4)%;(2.42±1.02 )% vs.(1 .23±0.42)%;(0.79±0.24)% vs.(0.26±0.11)%],the proportion of Breg cells was reduced significantly,the differ-ences had statistical significance[(0.83±0.21)% vs.(1 .89±0.12)%,P <0.05].In CITP children,the IFN-γ,IL-17,IL-22 levels in culture supernatant were higher than those in the control group[(278.2±121.2)pg/mL vs.(181.8 ±82.2)pg/mL;(214.8 ± 100.5)pg/mL vs.(161 .4±67.8)pg/mL;(122.16±22.2)pg/mL vs.(90.8±31.1)pg/mL],but the IL-10 level was lower than that in the healthy control group[(27.4±12.6)pg/mL vs.(46.1±16.2)pg/mL),differences between them had statistical signifi-cance(P <0.05).Besides,there was a positive correlation between the proportion of Breg cells and cytokine IL-10 level(r=0.617, P <0.05 ),however,there was a negative correlation between the proportion of Breg cells with Th1 ,Th17 and Th22 cells (P <0.05),between IL-10 level with IFN-γ,IL-17 and IL-22 levels(P <0.05).Conclusion The downregulation of Breg cells proportion may participate in the immune disorder mechanism of effective Th cells in CITP children,which can provide a new idea for the im-munoregulation therapy in CITP children.
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Objective To investigate the change of peripheral blood CD4+CD25+ regulatory T cells (Tr) in patients with idiopathic thrombocytopenic purpura (ITP), and to analyze its role in the pathogenesis of ITP.Methods Anticoagulated venous blood was collected from ITP patients (ITP group,n=35) and healthy controls (healthy control group,n=35). T lymphocytes were isolated and purified with human CD3+ T cell enrichment columm. The percentage of peripheral blood CD4+CD25+ regulatory T cells was detected by immunofluorescence staining (PE-anti-CD4 monoclone antibody and FITC-CD25 monoclone antibody) and bicolor flow cytometry by CD25/CD4 gating.Results The number and constituent ratio of CD4+CD25+T cell were significantly lower in ITP group than those of healthy control group (P<0.05).Conclusion There is peripheral blood celluar immunological function disorder in ITP patients, and decrease of CD4+CD25+ T cell population may be involved in the pathogenesis of ITP.