OBJECTIVETo develop a more reliable and stable method for determining monoclonal antibody (mAb) affinity constant based on competitive antibody/antigen binding.

METHODSThe Kd value was calculated based on the relationship between the binding proportion of the antigen and the original concentration of the mAb that competed for the binding site of the antigen.

RESULTSThe Kd values measured with this improved method under two different conditions were 2.61x10(-12) mol/L and 2.39x10(-12) mol/L, and those with Friguent method in these two conditions were 5.57x10(-10) mol/L and 1.41x10(-10) mol/L, respectively.

CONCLUSIONCompared with Friguent method, the Kd values measured with this improved method are closer to the actual value, and the measurement results under different experiment conditions are more stable.

Antibodies, Monoclonal ; immunology ; Antibody Affinity ; immunology ; Antigen-Antibody Complex ; immunology ; Binding, Competitive ; Enzyme-Linked Immunosorbent Assay ; Humans

To investigate leukocyte inhibitory factor(LlF) production and circulating immune complexes (CIC) in leprosy, peripheral blood mononuclear cells (PBMC) from 61 patients and sera from 6O patients were tested. The results indicate that there is a defect in LlF production in the lepromatous (LL) or borderline lepromatous (BL) types compared to the tubrculoid (TT) type (mean migration index=66.O +/- 16.O in LL 61.1 +/- 15.3 in BL, 51.9 +/- 11.2 in TT) (p < 0.05). The number of patients with positive CIC was higher among the LL patents (30%) than the TT patients (20%). There was also positive correlation between the bacterial index (Bl) and the CIC level (r=0.46, p < 0.05). The correlation between CIC and LIF in LL patients and the possibility (p=0.06) that the inuease m CIC may account for the decrease in LIF production in LL patients and vice versa are discussed.
Antigen-Antibody Complex/*analysis ; Human ; Leprosy/*immunology ; Leprosy, Lepromatous/immunology ; Leprosy, Tuberculoid/immunology ; Lymphokines/*analysis ; Support, Non-U.S. Gov't

BACKGROUNDSkin lesions are common manifestations in systemic lupus erythematosus (SLE). It is still unknown what the definite pathogenesis of skin involvement was and whether DNA participated in it. Our study was designed to explore the pathogenetic role and nature of nuclear antigen (DNA) deposited in the skin lesions of patients with SLE.

METHODSThirty skin samples from patients with SLE and 2 normal skin samples were studied. Extracellular DNA was evaluated by indirect immunofluorescence methods. The deposited immune complexes were extracted by cryoprecipitation, and DNA was then isolated with phenol and chloroform. DNA fragment sizes were detected by agarose gel electrophoresis. Finally, 8 different probes were used to analyze the origin of these DNA molecules using Dot hybridization.

RESULTSExtracellular DNA staining was found only in skin lesions, mainly those located in the basement membrane zone, vascular wall, and hair follicle wall. Normal skin and non-lesion SLE skin showed no fluorescence at locations outside the nuclei. There were no differences in the rate and intensity of extracellular DNA staining when comparing active phase to remission phase patients. No relationship was found between extracellular DNA and circulating anti-dsDNA antibodies. Deposited DNA fragments clustered into four bands of somewhat discrete sizes: 20 000 bp, 1300 bp, 800-900 bp, 100-200 bp. Small sized fragments (100-200 bp) were positively correlated with disease activity (P < 0.05, r = 0.407). Dot hybridization showed significant homology of the various extracellular DNA fragments examined with human genomic DNA, but not with DNA from the microorganisms and viruses we examined. There were also homologies between DNA samples from different individuals.

CONCLUSIONSDNA and its immune complexes may contribute to the pathogenesis of skin lesions in SLE. These DNA molecules range in size from 100 bp to 20 kb and may be endogenous in origin.

Antibodies, Antinuclear ; blood ; Antigen-Antibody Complex ; analysis ; DNA ; analysis ; immunology ; Humans ; Lupus Erythematosus, Systemic ; immunology ; Skin ; immunology ; Staining and Labeling

AIMTo explore the role of humoral immunity in the pathophysiological process of freezing injury and the possible immune interference in the preventation and treatment of frostbite.

METHODSSevere experimental freezing injury model was made in Wistar rats( n = 20). The concentration of three types of immunoglobulin (IgG, IgA and IgM), two types of complement components (C3 and C4), and circulating immune complex (CIC) were measured respectively before and at 4h, 1d, 3d, and 5d after frostbite. At the same time, the tissue immune complex (TIC) in skeletal muscle and the contents of the red blood cell immune complex (RBC-IC) were also observed and then was the red blood cell immune adherence activity (RCIA).

RESULTSSerum IgG concentration decreased rapidly to the lowest level at 4 h after frostbite IgA concentration dropped to the nadir on 1 day after freezing. Decreases of both immunoglobulins were maintained during the 5 days after frostbite. The fate of both C3 and C4 were the same as those immunoglobulins. Freezing had rather less effect on IgM level. CIC concentration in serum, expressed as the percent of prefreezing increased rapidly and to the zenith on the 3 days post-freezing. By immunofluorescence microscopy, thin continuous linear pattern (IgG) was demonstrated along the SM on the first day post-freezing. Granular and nodular deposits (IgG) appeared along the SM as the time proceeded after frostbite. RBC-IC contents, expressed as the erythrocyte IC rosette rate, increased significantly and to the zenith on the 3 d post-freezing, while RCIA depressed to the nadir at the same time.

CONCLUSIONThe freezing frostbite is an immune complex related disease which have not been reported by others before.

Animals ; Antigen-Antibody Complex ; analysis ; immunology ; Frostbite ; blood ; immunology ; Immunoglobulin A ; immunology ; Immunoglobulin G ; immunology ; Immunoglobulin M ; immunology ; Immunoglobulins ; immunology ; Male ; Rats ; Rats, Wistar

Higher levels of IgE-containing immune complexes (IC) have been reported in sera from patients with allergic diseases than in sera from controls. To evaluate the possibility of an IC-mediated mechanism in the pathogenesis of bronchial asthma, we measured circulating C3-containing IgE IC (C3-IgE IC) using anti-C3 ELISA from 20 house dust mite (HDM)-sensitive asthmatics, 20 non-atopic asthmatics, and 14 non-atopic controls. C3-IgE IC levels were significantly higher in HDM-sensitive asthmatics (mean +/- S.D.: 12.2 +/- 7.8 AU/ml) than in non-atopic asthmatics (6.5 +/- 7.5 AU/ml) or controls (5.8 +/- 4.4 AU/ml). C3-IgE IC levels were significantly correlated with HDM-specific IgE levels (r = 0.50, p<0.05), but not with total IgE levels (r = 0.36, p< 0.05) in HDM-sensitive atopic asthmatics. C3-IgE IC levels in sera did not significantly change during HDM-bronchoprovocation test in six HDM-sensitive asthmatics who showed positive reaction. Part of C3-IgE IC could be precipitated by protein G coupled beads. In conclusion, C3-IgE IC levels were elevated in sera from HDM-sensitive asthmatics; moreover IgG antibodies might be a component of C3-IgE IC. Our results suggest that an IgE IC-mediated mechanism could be involved in the pathogenesis of atopic asthma.
Adult ; Animal ; Antigen-Antibody Complex/*blood ; Asthma/*immunology ; Complement 3/*analysis ; Dust ; Human ; Immunoglobulin E/*blood ; Mites/immunology

Heparin-induced thrombocytopenia (HIT) is an antibody-mediated complication of heparin treatment that can lead to thrombosis and thromboembolism. HIT is mainly caused by immunoglobulin G (IgG) class among anti-heparin/platelet factor 4 antibodies that bind to epitopes on platelet factor 4 (PF4) released from activated platelets that developed when it forms complexes with heparin. Platelet aggregation and hypercoagulation status result from this process. Besides, the reactions between antibodies and vascular endothelial cells and monocytes are involved in HIT. Laboratory detection of anti-heparin/platelet factor 4 antibodies after heparin administration may help diagnose HIT early. Tests for detecting antibodies to the heparin/PF4 complex can be classified into functional platelet assays (which rely on the demonstration of platelet activation) and immunoassays (which detect the presence of an antibody without regard for its functional ability). But there is no simple and effective test available currently. In this article the anti-heparin/platelet factor 4 antibodies, pathogenesis of HIT, clinical laboratory assays and immunoassays are reviewed.
Antigen-Antibody Complex ; immunology ; Heparin ; adverse effects ; immunology ; Humans ; Immunoglobulin G ; immunology ; Platelet Aggregation ; Platelet Factor 4 ; immunology ; Thrombocytopenia ; chemically induced ; immunology

OBJECTIVETo explore the characteristics of Helicobacter pylori (H. pylori) antigen in serum and to evaluate its clinical diagnostic value.

METHODSEnzyme-linked immunosorbant assay (ELISA) was developed to detect the soluble H. pylori antigen (S-Hp) and circulatory specific H. pylori antigen immunocomplexes (Hp-IC) in serum.

RESULTSThe positive rate of S-Hp was 90.91% from 66 patients with H. pylori infection, which was much greater than 0% found in 28 controls (P < 0.001). Moreover, its concentration closely reflected the number of H. pylori in the gastric mucosa layer. We also found that Hp-IC existed bound with IgG and/or IgA in patients with positive S-Hp. However, there is no evidence to show the concentration of S-Hp reduced significantly in followed-up subjects after effective therapy.

CONCLUSIONSThese methods as newly and noninvasive complementary tools can be used for clinical diagnosis of H. pylori infection. In addition, S-Hp and Hp-IC may be of importance in H. pylori pathogenesis.

Adult ; Antibody Specificity ; Antigen-Antibody Complex ; blood ; Antigens, Bacterial ; blood ; Female ; Helicobacter Infections ; immunology ; Helicobacter pylori ; immunology ; Humans ; Immunoglobulin A ; blood ; Immunoglobulin G ; blood ; Male

OBJECTIVETo study the correlation of erythrocyte immune function between normal neonates and their mothers and the influence of various obstetric factors on neonatal erythrocyte immune function.

METHODSThe adherent rate of complement 3b-receptor on the surface of red blood cells (RBC-C3bRR) and the immune complex adherent rate of red blood cells (RBC-ICR) were detected using the erythrocyte saccharomyces rosette test in 104 normal neonates and their mothers. The correlation of erythrocyte immune function between neonates and their mothers was evaluated by the maternal-infant paired test.

RESULTSThe levels of RBC-C3bRR (16.80 +/- 1.56% vs 16.23 +/- 1.63%; P < 0.05) and RBC-ICR (5.72 +/- 1.63% vs 5.02 +/- 1.38%; P < 0.01) in neonates were significantly higher than those in their mothers. There was a significantly positive correlation in RBC-ICR levels between neonates and their mothers (r = 0.28, P < 0.05). No correlation was found in RBC-C3bRR levels between the two groups. Neither RBC-C3bRR nor RBC-ICR levels of neonates were associated with various obstetric factors such as amniotic fluid, placenta, umbilical cord, parturient patterns, and puerperal anemia and pregnancy-induced hypertension syndrome.

CONCLUSIONSThe erythrocyte immune function in neonates has a relatively mature level and correlates with their mothers' erythrocyte immune function. Various obstetric factors have no influences on neonatal erythrocyte immune function.

Antigen-Antibody Complex ; immunology ; Erythrocytes ; immunology ; Female ; Fetal Blood ; immunology ; Humans ; Infant, Newborn ; immunology ; Linear Models ; Male ; Pregnancy ; Receptors, Complement 3b ; analysis ; Rosette Formation

The occurrence of immune complexes in the serum from rats infected with M. leprae-murium and 38 patients with leprosy were studied by the polyethylene glycol precipitation complement consumption (PEG-CC) test and the results were compared in the various forms of the disease. Circulating immune complexes (CIC) were significantly increased in the sera from rats infected with M. lepraemurium compared to normal control rats (P < 0.005). There were no significant differences between the the level of CIC in the sera from lepromatous leprosy patients and that from tuberculoid leprosy patients, but in the sera from patients with erythema nodosum leprosum (ENL) the level of CIC was significantly increased (P < 0.005). And although we couldn't find a corre1ation between the level of CIC and bacterial indices in lepromatous leprosy patients, CIC tends to de-crease after negative conversion of their bacterial indices. These findings suggested that the detection of CIC can be of some practical interest in the early diagnosis of ENL and can be a valuable assessment in following the therapy after negative conversion of their bacterial indices.
Animal ; Antigen-Antibody Complex/analysis* ; Disease Models, Animal ; Human ; Leprosy/immunology* ; Male ; Rats ; Reference Values ; Time Factors

Nonspecific immune parameters such as natural killer(NK) activity, antibody-dependent cellular cytotoxicity(ADCC), production of leukocyte migration inhibitory factor(LlF) and levels of immune complex(IC) were assessed in 47 patients with rheumatoid arthritis (RA) 20 with degenerative arthritis (DA) and 40 healthy controls. Peripheral blood (PB) as well as synovial fluid (SF) were collected from both RA and DA patients before treatment. Mononuclear cell suspensions and sera were prepared and submitted for the in vitro tests; 4-hr chromium-release assays using human K562 and mouse L1210 cells as targets for NK and ADCC assays respectively, 2-step agarose assay for LIF and platelet aggregation test for IC. Results revealed that 1) LIF activity of PB lymphocytes (PBL) from both RA and DA patients showed a significant (P < 0.05) decrease as compared with that from healthy controls. 2) PB-NK activity from RA patients showed an insignificant decrease as compared with that from DA or healthy controls. However, mononuclear cells isolated from SF (SFL) of RA patients exhibited significantly(P < 0.02) lower NK activity than PBL from the same patients. 3) In ADCC assays with PBL no significant differencies were observed among the 3 groups. 4) Higher titers of IC were detected in both PB and SF from RA patients than DA, and a negative correlation was found between serum IC levels and PB-NK activity. These data are discussed in light of previous reports, and a hypothesis regarding a decreased nonspecific cell-mediated immunity in conjunction with an increased humoral immune response, particularly in local sites, is proposed as one of the mechanisms underlying the pathogenesis of RA.
Adult ; Antibody-Dependent Cell Cytotoxicity ; Antigen-Antibody Complex/immunology ; Arthritis, Rheumatoid/immunology* ; Female ; Human ; Killer Cells, Natural/immunology ; Leukocyte Migration-Inhibitory Factors/biosynthesis ; Male ; Middle Age ; Synovial Fluid/immunology