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Objective:To establish a rapid and quantitative method for the determination of immunoglobulin G4 (IgG4) by fluorescence immunochromatography and to analyze its clinical application value.Methods:Fluorescence immunoassay for quantitative detection of IgG4 was obtained by means of preparation of kits in a competitive reaction mode and combining immunoassay with fluorescence quantitative assay. The linearity, precision, accuracy, anti-interference ability and stability of the method were evaluated, and compared with immune-scattering turbidimetry, receiver operating characteristic curve (ROC) was plotted, area under the curve (AUC) was calculated, and the critical value for the diagnosis of pancreatitis related diseases was determined, and sensitivity and specificity were calculated.Results:The linear range of fluorescence immunoassay for IgG4 was 0.2-10.0 g/L. The accuracy coefficient of variation was less than 15%, and the accuracy deviation was within ±15%. Bilirubin (2.5 g/L), triglyceride (10 g/L) and hemoglobin (10 g/L) had no significant effect on the quantitative determination. Within 14 months, 1.20 g/L and 2.65 g/L reference samples were detected with concentration deviations within ±15%. The kit validity period was >12 months. Serum samples of 200 healthy people were detected by fluorescence immunochromatography, and the normal reference value of IgG4 was <2.03 g/L. fluorescence immunochromatography and Immunoturbidimetry were used to detect IgG4 concentrations in 383 clinical serum samples. The results showed that the two methods were consistent ( P>0.05). Using 2.01g/L IgG4 as the critical value, the sensitivity and specificity of fluorescence immunochromatography were 96.3% and 95.5% by ROC curve analysis, respectively. Conclusions:Fluorescence immunochromatography was a simple, rapid and accurate method for the quantitative detection of IgG4, and had high sensitivity and specificity for the diagnosis of pancreatitis related diseases. It was suitable for quantitative detection of bulk samples in outpatient and emergency departments.
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Objective To conduct a methodological evaluation on flowcytomotry fluorescence detection of serum CA242, and to evaluate its value in the diagnosis of pancreatic cancer.Methods The blood samples of 40 cases of pancreatic cancer, 49 cases of other tumors, 48 cases of benign digestive diseases and 159 healthy volunteers were collected.Fowcytometry fluorescence immunoassay was used to detect serum level of CA242, and it was compared with routine ELISA method to measure its sensitivity and specificity.Results The detection limit of CA242 by flowcytometry fluorescence immunoassay was 0.89 U/ml, the linear range was 1～ 500 U/ml after confirmation.The within-batch CV was 3.37％～ 5.30％, between batch CV was 7.43％ ～ 9.60％.When compared with routine ELISA, flowcytometry fluorescence immunoassay showed the equation of linear regression is Y =1.0398X-0.947, r =0.9687.Area under ROC curve was 0.811 ± 0.025 (95％ CI 0.763 ～0.859), with 18.625 U/ml as the best cutoff value, the specificity was 92.0％ and the sensitivity was 62.1％.Conclusions Flowcytometry fluorescence immunoassay for CA242 testing has the advantage of shorter detection time, miltiple sample and testing project detection, which is worth of clinical application.
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Objective To study the regulatory effects of Ca 2+ , ATP, ADP and AMP on myocardial nuclear Ca 2+ ATPase activity after ischemia reperfusion injury Methods The model of myocardial ischemia reperfusion injury was established in rats Myocardial nuclei were purified using sucrose density gradient centrifugation The activity of Ca 2+ ATPase was measured with ascertaining phosphate Results Compared with the control, plasma levels of MDA and FFA after myocardial ischemia reperfusion injury increased significantly ( P
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Aim Observing the alteration of cardiac myocyte nuclear inositol 1,4,5-trisphosphate receptor (IP_3R)binding proterties in rat subjected to myocardium ischemic reperfusion is to make it clear whether this change is involved in the molecule mechanism of cell apoptosis of rat with myocardial ischemic reperfusion. Method Apoptosis index of myocardial cell was determined using TUNEL assay.Extracting of cardiac myocyte nucleus was accomplished by saccharose density gradient centrifugation method,the binding proterties of nuclear IP_3R in different conditions were detected by radioligand binding assay.Results ①Myocardial cell apoptosis index in rat heart underwent 30 min regional ischemia and 3 h reperfusion was distinctly increased compared with sham-operated group(P
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AIM: To investigate the relationship between serum soluble E-selectin (sE-selectin) and insulin resistance, serum uric acid, serum lipid in essential hypertension patients. METHODS: Fasting serum sE-selectin concentration, plasma glucose, serum insulin, serum uric acid, total cholesterol, triglycerides, high density lipoprotein-cholesterol, low density lipoprotein-cholesterol were determined in 186 patients with essential hypertension (75 males, 111 females). Homeostasis model assessment was applied to assess the status of insulin resistance (HOMA-IR). RESULTS: Based on the HOMA-IR, the essential hypertension patients were divided into insulin-sensitive individuals (IS) and insulin resistant subjects (IR). The serum sE-selectin concentration was significantly higher in male group [(50.1?17.8)?g/L] than in female group [(40.6?16.6)?g/L] (P
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AIM: The changes of myocardial nuclear membrane Ca 2+ -ATPase function was investigated in ischemia/reperfusion injury. METHODS: The model of myocardial ischemia/reperfusion injury was established in rats. Myocardial nuclei were purified with sucrose density centrifugation,the activity of Ca 2+ -ATPase was measured and calcium uptake was assayed with [ 45 Ca 2+ ] . RESULTS: Plasma levels of malondialdehyde (MDA) and free fatty acid (FFA) in myocardial ischemia/reperfusion injury increased significantly( P
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Objective To study the relationship between innate immunity of lymphocytes and hu-moral and cell-mediated immunity in patients with psoriasis.Methods The rapid natural immune reaction on cancer cells was measured by lymphocytes isolated from fresh blood.Serum IgG,IgA,IgM were measured with rate-nephelometry in patients with psoriasis,and sandwich ELISA technique was applied to detect serum sIL-2R level in those patients.Results Rosette formation rate of lymphocytes adhering to cancer cells and serum immunoglobulin IgA level were significantly higher in patients with psoriasis than those in normal controls(P
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Objective To study CD35on erythrocytes and erythrocyte chemokine recepter(ECKR)in patients with psoriasis in order to explore red blood cell(RBC)innate immune function and its possible role in the pathogenesis of psoriasis.Methods The rapid natural immune reaction on cancer cells was measured with RBCs isolated from fresh blood.Expression of CD35on erythrocytes was detected by flow cy-tometry.The level of IL-8,which was bound to isolated erythrocytes,was measured by enzyme linked im-munosorbent assay(ELISA)in the supernatant after centrifugation,which represented ECKR binding activity.Results Rosette formation rates of RBC adhering to cancer cells and expression of CD35on RBCs were significantly higher in psoriatic patients than those in control group(P
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AIM: To investigate the changes in nuclear calcium content and permeability of nuclear pore complex in rat myocardium during ischemia reperfusion injury.METHODS: The rat model of myocardial ischemia reperfusion injury was established.Myocardial nuclei were purified using sucrose density gradient centrifugation.The nuclear calcium content was measured by atomic absorption spectrophotometer.The permeability of nuclear pore complex was assessed through measuring the amount of calmodulin conjugated Alexa Fluo~(TM) 488 as fluorescent probes transported across nuclear membrane with spectrofluorometer.RESULTS: The nuclear calcium content at 15,30,60,120 and 180 min reperfusion following 30 min sustained ischemia increased 1.31-,1.55-,1.73-,1.94-and 2.14-fold,respectively,as compared with sham-operation group.The permeability of nuclear pore complex at 15 min reperfusion following 30 min sustained ischemia showed no difference from sham-operation group,but it only increased 1.31-,1.38-,1.40-,and 1.48-fold at 30,60,120 and 180 min reperfusion following 30 min sustained ischemia compared with sham-operation group.CONCLUSION: The nuclear calcium content during myocardial ischemia reperfusion injury increases earlier than the permeability of nuclear pore complex does.The increase in the permeability of nuclear pore complex may result in adaptive regulatory effects on nuclear calcium overload to a certain extent during myocardial ischemia reperfusion injury.
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Colorimeter detection of Evas Blue in formamide extract of rat tracheal tissue is used as an indication of increased vascular permeability induced by introduction of 115 degrees C air into the rat trachea, followed by a dose (20mg/kg) of meperidine given. The increases of tracheal vascular permeability (TVP) and malondialdehyde (MDA) level induced by hot air injury (HAI) are markedly inhibited. The decrease of superoxide dismutase (SOD) activity induced by HAI is raised. The inhibitory effect of meperidine (MIE) on TVP is not antagonized by naloxone. It is implied that the opiate receptor sensitive to naloxone is not involved in the MIE on TVP. The decrease of tracheal MDA content, increase of tracheal SOD activity raising the ability of scavengering oxygen free radicals and reducing the formation of other active oxygen, inhibit lipid peroxidation. These antioxidative effects may be one aspect of the mechanism of the inhibiory effect of meperidine on TVP.
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Objective To evaluate of red blood cell as giving instruction in whole white blood cell immunological activity by new nature experimental system of hemaimmune reaction rood map. Methods Plasma 0.3ml were added to whole blood cells (including: red blood cell and white blood cells) or white blood cells 0.2ml, and incubated for 1h at 37℃. The content of IL-8 and IL-12 was determined by enzyme linked immunadsorbent assay (ELISA) method. The expression level of CD4, CD8, CD35 and CXCR4 on white blood cells was determined by method of Flow Cytometry. Results The content of IL-8 (5.96?4.26) and IL-12 (9.84?2.23) in whole blood and plasma nature group was significantly lower than that (13.59?3.69?B?pg~ -1 ?ml~ -1 ) and (15.09?9.86?B?pg~ -1 ?ml~ -1 ) in white blood cell and plasma isolation group (P
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Objective To determine the effect of antigen on the main immunological reaction route of red blood cells and while blood cells. Methods Cancer cells (5?10~6/ml) and/or Bacillus calmette-Guerin(BCG 0.1mg) or yeast cells(5?10~8/ml) were added into 0.2ml of whole blood cells (or 0.2ml of white blood cells) and 0.3ml of fresh plasma (or 0.3ml of NS) treated by citric acid, and incubated for 1h at 37℃. IL-8 level was measured by ELASA. The data could be divided into 4 groups. (1) 0.2ml of antigen (cancer cells or yeast cells or BCG) was added to 0.2ml of whole blood cells and 0.3m plasma. ②0.2ml of NS was added to 0.2ml of whole blood cells and 0.3ml of plasma. ③ 0.2ml of antigen was added to 0.2ml of white blood cells and 0.3ml of plasma. ④ 0.2ml of NS was added to 0.2ml of white blood cells and 0.3ml of plasma. Results Cancer cells, BCG and yeast cells could activate immunological reaction in blood, but could not activate immunological reaction of white blood cells in no plasma group with addition of antigen. The activation Index (2.124?0.860) of IL-8 in the group with addition of whole blood cells and plasma was significantly higher than that (0.390?0.08) in the group with addition of antigen, white blood cells and plasma (P
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Objective To approach the capacity of erythrocyte regulating IL-8 in patients with primary hepatocarcinoma (PHC) by a new experimental system of hemaimmune. Methods 0.2ml suspension of cancer cells (S180: 5?10~6/ml) or NS were added into 0.2ml anticoagulant suspension of whole blood cells or leukocytes and 0.3ml plasma, then incubated for 1 h at 37℃. The content of IL-8 was determined by ELISA. Results In the patients with PHC, the IL-8 levels (pg/ml) in experimental and control groups of whole blood cells, and in experimental and control groups of leukocytes were 376.35?243.96, 353.64?271.92, 461.27?277.11 and 424.97?278.93, respectively; while in the normal human, they were 11.36?6.93, 4.98?4.35, 29.41?30.66 and 20.77?24.20, respectively. In the patients with PHC, the activation rates of cancer cells in the experimental groups of both whole blood cell and leukocyte were 0.22?0.24 and 0.25?0.53, respectively; while in normal human, they were 2.49?2.33 and 0.75?0.21, respectively. In the patients with PHC, the IL-8 adsorption rate of erythrocyte in both experimental and control groups of whole blood cell were 0.22?0.18 and 0.17?0.33, respectively; while in normal human, they were 1.18?2.29 and 0.86?0.49, respectively. The IL-8 activated rates of erythrocyte in the patients with PHC was much lower than in the normal human (P
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Objective To study the effect of red blood cells on the secretion of white blood cells modulating globulin in patients with cancer with a new immunological reaction experimental system. Methods Cancer cells (S180, 5?10~6/ml) or 0.2ml of NS or 0.2ml of white blood cells and 0.3ml of plasma were added to anticoagulant suspension of white blood cells treated by citric acid and incubated for 1h at 37℃. The content of globulin (g/L) was determined by Olympus AU100 detector method. 0.2ml of cancer cells was added to 0.2ml of whole blood cells and 0.3ml of plasma in the whole blood experimental group. 0.2ml of NS was added to 0.2ml of whole blood cells and 0.3ml of plasma in the control group. 0.2ml of cancer cells was added to 0.2ml of white blood cells and 0.3ml of plasma in the white blood cell isolation experimental group. 0.2ml of NS was added to 0.2ml of white blood cells and 0.3ml of plasma in the control group. Results Activation rate (0.31?0.09) of globulin in whole blood cell experimental group of patients with cancer was significantly decreased compared to that (0.39?0.14) in normal people. Red cell adhesion rate (0.09?0.08) of globulin in control group was also significantly decreased compared to that (0.29?0.14) in normal people. Conclusion The results of our study indicate that the new experimental system can be used for detection of red blood cell-modulating globulin. The function of red blood cells in modulating globulin of patients with cancer is decreased.
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Objective:To investigate the feasibility of treating chronic HBV infection by inducing spe cific humoral immune response to HBV middle envelope protiens in normal and HBV transgenic mice with HBV DNA based immunization. Methods: The eukaryotic expression vector pCMV S2.S (PS) containing HBV S2.S gene and pc DNA3.0 were used respectively to immunize 5 C57BL/6 normal mice and HBV transgen ic mice. Each mouse was injected intramuscularly with one of those plasmids at t he same dose (100 ?g). Sera of mice were detected for anti HBs, anti preS2, HBsAg and HBeAg with ELISA. Pathological changes of transgenic mice liver were o bserved by microscopy. Results:PS can stimulate immune respons es of anti HBs and anti preS2 in normal and transgenic mice.The appearance of a nti preS2 was 1 2 weeks earlier than that of anti HBs. HBsAg and HBeAg in se ra turned negative 8 weeks after immunization. At the 8th week, hepatocytes show ed extensive granular degeneration and hydropic degeneration. There was no obvious difference in the amount of mononuclear lymphocytes between pre and post gen e immunization. Conclusion: It is showed that specific humoral immune response can be effectively induced by PS after DNA based immunization, and PS seems to be responsible for the disapearance of HBsAg and HBeAg in sera o f HBV transgenic mice. The results provide an evidence for furth er investigation of genetic vaccine in the treatment of chronic HBV infection.
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Objective To study the change in the rule of quick innate immune reactivity of fresh blood corpuscles to cells antigen. Methods Microqrganisms ( Yeast , Escherichia coli , Pseudomanas aeruginosa ) were added in blood anti coagulated by citric sodium and incubated at 37℃ for 30min. Results The innate immune reactivity of the red blood corpuscles to yeast was significantly higher than that of white blood cells ( P