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Objective:To detect serum levels of immunoglobulin E (IgE), soluble version of the IgE-binding alpha-chain of Fcepsilon-RI (sFcεRIα), immunoglobulin G (IgG) anti-IgE, IgG anti-FcεRI in patients with chronic urticaria(CU), and to evaluate their relevanceMethods:Forty-three newly diagnosed patient with CU were enrolled. Thirty-seven patients with dermatitis or eczema and 51 healthy subjects were selected as the disease control (DC) and normal control (NC) group, respectively.Serum IgE was detected by immunoturbidimetry; serum anti-IgE, anti-FcεR Ⅰ and sFcεR Ⅰ α were determined byenzyme-linked immunosorbent assay (ELISA) methods. Statistical analysis was carried out by non-parametric test for comparisons of the above variables between groups, and by Spearman correlation analysis for assessment of relationships between the variables as well as between the variables and disease severity, and by receiver operating characteristic curve to analyze the diagnostic value of the variables.Results:Serum IgE, anti-IgE and anti-FcεRI in CU were significantly higher than that of NC.Their medians are 65.70 IU/ml vs 17.10 IU/ml,χ 2=28.541, P=0.001;0.61 vs 0.39,χ 2=27.408, P=0.001;0.64 vs 0.51, χ 2=29.102, P<0.001.Serumanti-FcεRI in CU was significantly lower than that in NC(0.64 vs 0.83,χ 2=25.869, P=0.007).The medians of serum sFcεRIα in CU, DC and NC groups were 5.74,5.38,4.50 ng/ml, respectively. The difference was not statistically significant,χ 2=3.463, P=0.177.There was a positive correlation between IgE and sFcεR Ⅰ α, anti-IgE and anti-FcεRI( r=0.455, P<0.001; r=0.611, P<0.001).No significant correlation was showed between the four variables and the course of disease or the severity of symptoms, P>0.05. The diagnostic performances of IgE, anti-FcεRI for CU were similar (AUC=0.72),which were better than that of sFcεRIα (AUC=0.61).The highest diagnostic efficacy (AUC=0.83) can be achieved by four joint tests.Anti-FcεRI is of value in differential diagnosis of CU and DC, AUC=0.7, P=0.002. Conclusion:The levels of serum IgE, anti-IgE, anti-FcεRI were significantly increased in CU patients, and these mast cell activation-related molecules have the potential to be diagnostic markers for CU.
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Objective@#The accurate measurement of anti-IgE levels in patients′ serum helps for make diagnosis of chronic urticaria (CU). An indirect ELISA method for quantitative detection of IgG anti-IgE, were established.@*Methods@#The serum samples and the clinical data of 75 first-diagnosed CU patients and 120 healthy controls were collected at Shanghai General Hospital during the year of 2018. In the indirect ELISA system to measure the IgG (anti-human IgE), the antigen, Human IgE Myeloma, was coated on the plate; Omalizumab (a humanized anti-human IgE antibody) was the standard, and horse radish peroxidase (HRP)-labeled anti-human IgG was the tracer. The optimum concentrations of serum and HRP-labeled anti-human IgG were determined by chessboard titration, and method evaluation was conducted. The comparison of serum anti-IgE levels in CU patients and healthy people were analyzed by Mann-Whitney U test and chi-square test. Receiver operating characteristic (ROC) curves were applied to establish the diagnostic performance of serum anti-IgE for CU.@*Results@#The coating concentration of IgE was 5.0×10-4 mg/ml; serum dilution was 1∶300; enzyme-labeled antibody dilution was 1∶100 000. Standard curve was in good linearity with R2=0.996. The intra-and inter-assay coefficient of variation were 3.9%-7.5% and 6.0%-8.2%, and the recovery rate of low and high concentration samples were 95.9% and 108.4%, respectively. When hemoglobin≤1.3 g/L, triglyceride≤4.6 mmol/L, bilirubin≤171 μmol/L, no interference were observed. The limit of blank, limit of detection, and limit of quantitation were 5.8×10-4, 1.8×10-3, and 2.0×10-3 mg/ml. The linearity range was from 2.0×10-3 to 354.4 mg/ml. No Hook effect was found until anti-IgE reached 354.4 mg/ml. The anti-IgE remained stable after serum storage at 4 ℃ overnight or treated with 6 repeated freeze-thaw cycles. The anti-IgE levels in CU patients [19.0(1.9, 58.6)mg/ml] were significantly higher than those in healthy controls [0.7(0.002, 11.1)mg/ml] with P<0.001. When cut-off value was set as 15.3 mg/ml, in this method, the positive rate of CU patients (54.7%) was significantly higher than these of healthy controls (18.3%) (P<0.001, AUC=0.736).@*Conclusions@#The indirect ELISA method for serum anti-IgE measurement was successfully established. Anti-IgE autoantibodies in serum would be used in the diagnosis of CU.
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Objective The accurate measurement of anti-IgE levels in patients' serum helps for make diagnosis of chronic urticaria (CU). An indirect ELISA method for quantitative detection of IgG anti-IgE, were established. Methods The serum samples and the clinical data of 75 first-diagnosed CU patients and 120 healthy controls were collected at Shanghai General Hospital during the year of 2018. In the indirect ELISA system to measure the IgG (anti-human IgE), the antigen, Human IgE Myeloma, was coated on the plate;Omalizumab (a humanized anti-human IgE antibody) was the standard, and horse radish peroxidase (HRP)-labeled anti-human IgG was the tracer. The optimum concentrations of serum and HRP-labeled anti-human IgG were determined by chessboard titration, and method evaluation was conducted. The comparison of serum anti-IgE levels in CU patients and healthy people were analyzed by Mann-Whitney U test and chi-square test. Receiver operating characteristic (ROC) curves were applied to establish the diagnostic performance of serum anti-IgE for CU. Results The coating concentration of IgEwas 5.0 × 10-4 mg/ml; serum dilution was 1:300; enzyme-labeled antibody dilution was 1:100000. Standard curve was in good linearity with R2=0.996. The intra-and inter-assay coefficient of variation were 3.9％-7.5％ and 6.0％-8.2％, and the recovery rate of low and high concentration samples were 95.9％ and 108.4％, respectively. When hemoglobin≤1.3 g/L, triglyceride≤4.6 mmol/L, bilirubin≤171μmol/L, no interference were observed. The limit of blank, limit of detection, and limit of quantitation were 5.8 × 10-4, 1.8 × 10-3, and 2.0 × 10-3 mg/ml. The linearity range was from 2.0 × 10-3 to 354.4 mg/ml. No Hook effect was found until anti-IgE reached 354.4 mg/ml. The anti-IgE remained stable after serum storage at 4℃overnight or treated with 6 repeated freeze-thaw cycles. The anti-IgE levels in CU patients [19.0(1.9, 58.6)mg/ml] were significantly higher than those in healthy controls [0.7(0.002, 11.1)mg/ml] with P<0.001. When cut-off value was set as 15.3 mg/ml, in this method, the positive rate of CU patients (54.7％) was significantly higher than these of healthy controls (18.3％) (P<0.001, AUC=0.736). Conclusions The indirect ELISA method for serum anti-IgE measurement was successfully established. Anti-IgE autoantibodies in serum would be used in the diagnosis of CU.
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Objective:To investigate the activation of peripheral basophils from patients with rheumatoid arthritis ( RA) and its mechanisms. Methods:The activation markers including CD203c and the proportion of IL-4 positive rate of peripheral basophils from RA patients and healthy controls were detected by flow cytometry. Serum levels of IgE in RA patients and healthy controls were detected by electrochemilu minescence immunoassay. Basophils were negatively isolated and co-cultured with or without purified IgE,anti-IgE as positive control,then,the expression of CD203c and propotion of IL-4 positive basophils were detected by flow cytometry. Results:The expression of CD203c and IL-4 positive rate of basophils from RA patients were higher than that of healthy controls (P<0. 05). Serum levels of IgE in RA were higher than that of healthy controls (P<0. 05). After co-cultured with isolated IgE from RA patients,basophils negatively isolated from healthy controls were activated,and higher expression of CD203c and proportion of IL-4 (P<0.05). Conclusion:Basophil activation is related with development of RA,and its activation is mainly mediated by IgE. Targeting basophil and its activation pathway would be expected to provide new strategies for the treatment of RA.