Sera Of Control Subjects Research Articles

Autoantibody Profile in Systemic Lupus Erythematosus Patients

Systemic lupus erythematosus (SLE) is an autoimmune disease, in which the etiology is not well-understood; however, interactions between environmental and genetic factors in predisposed individuals have been recognized. As a consequence, immunological alternations occur and immune cells are involved, especially T and B lymphocytes that are activated to produce different immune components. Among these components are autoantibodies that react with self-antigens aside from non-self-antigens due to the proposed theory of molecular mimicry. Accordingly, the current study was designed to examine the profile of different autoantibodies in SLE patients by using the indirect membrane based enzyme immunoassay for the quantitative measurement of IgG class antibodies. Subjects: Sixty-four SLE patients (32 arthritis and 32 nephritis patients) and 32 healthy subjects (control) were enrolled in the study, and their sera were tested for anti-nucleosome, anti-histone, anti-smD1, anti-PCNA, anti-PO, anti-SS-A/Ro-60, anti-SS-A/Ro-52, anti-SS-B/La, anti-CENP, anti-SCI-70, anti-U1snRNP, anti-AMA-M2, anti-Jo-1, anti-PM-SCI, anti-Mi2 and anti-Ku autoantibodies in order to evaluate the autoimmunity status in SLE patients. Results: The sera of control subjects were negative for these antibodies; therefore, the comparisons were limited to the two groups of SLE patients; arthritis and nephritis. The highest percentage of seropositive arthritis patients was observed for anti-SS-A/Ro-60, anti-CENP and anti-U1snRNP antibodies (100.0%), while the lowest percentage was recorded for anti-Jo-1 antibody (15.6%). For nephritis patients, anti-U1snRNP antibody (87.5%) was also observed to have the highest percentage, and anti-Jo-1 antibody (3.1%) also recoded the lowest percentage. However, four autoantibodies (anti-PCNA, anti-SS-A/Ro-60, anti-SS-B/La and anti-CENP antibodies) showed different profiles in arthritis and nephritis SLE patients. They showed a significant increased percentage in arthritis patients compared to nephritis patients (anti-PCNA: 87.5 vs. 50.0%, p = 0.003; anti-SS-A/Ro-60: 100.0 vs. 81.2%, p = 0.02; anti-SS-B/La: 75.0 vs. 43.8%, p = 0.02; anti-CENP: 100.0 vs. 43.8%, p = 0.001). Conclusion: These findings suggest the diagnostic potential of autoantibodies as early markers for SLE development.

ASSOCIATION OF INTERLEUKIN-4 CYTOKINE AND IL-4Rα GENE POLYMORPHISM IN β-LACTAM ALLERGIC PATIENTS

Objective: The present study was carried out to estimate the possible role of Interleukin-4 (IL-4)RαQ576R genes polymorphism in the development of immune reaction against penicillin, as well as to study the effect of IL-4 cytokine in regulating allergic reactions.Materials and Methods: Measurement of serum IL-4 concentration was done using enzyme-linked immunosorbent assay technique; IL-4RαQ576R gene polymorphisms were genotyped using polymerase chain reaction-restriction fragment lengths polymorphisms. Comparisons for statistical significance were performed using Mann–Whitney U-test.Results: Comparing with control subjects, there was a significantly increased level of IL-4 (348.53 pg/ml) in penicillin allergic patients versus (284.72 pg/ml) in sera of control subjects. The IL-4RαQ576R alleles were significantly higher in the penicillin allergic individual compared with apparently healthy control subjects.Conclusions: Data study suggested that IL-4 cytokine have some important roles in penicillin hypersensitivity reaction, additionally the IL- 4RαQ576Rgene polymorphisms might involve in modulating of penicillin hypersensitivity.

ELISA for human serum leucine-rich alpha-2-glycoprotein-1 employing cytochrome c as the capturing ligand

Leucine-rich alpha-2-glycoprotein-1 (LRG) is a serum glycoprotein of unknown function that has shown promise based on qualitative assessments as a biomarker for certain diseases including microbial infections and cancer. However, the lack of a quantitative assay for LRG has limited its application. Here an indirect enzyme-linked immunosorbent assay (ELISA) for quantifying LRG in human serum is described in which cytochrome c is employed as the capturing ligand and a monoclonal antibody specific for LRG is used to detect the captured glycoprotein. Application of this assay in quantifying LRG in various patients' sera is demonstrated. The concentration of LRG in sera of control subjects as determined by this assay is approximately 50 μg/ml. Consistent with expectations from published reports, LRG was found to be significantly elevated in the sera of some patients with a bacterial infection (toxic shock syndrome, TSS). LRG was only slightly elevated in patients infected with the human immunodeficiency virus as compared to uninfected control subjects, while normal levels of LRG were observed in patients with non-infectious diseases (inflammatory arthritis and neurological disorders, primarily Parkinson's disease). Although LRG and C-reactive protein (CRP) are both produced by the liver and are classified as acute-phase proteins, there was no significant correlation between the levels of LRG and CRP in the sera of the patients. Thus, LRG and CRP measurements are non-redundant and indicate different physiological contexts. The ELISA described in this report should be useful to further assess serum LRG as a biomarker for clinical diagnostics.

Serum protein MALDI profiling to distinguish upper aerodigestive tract cancer patients from control subjects.

There are no reliable blood markers for the early detection and monitoring of aerodigestive tract tumors. Recent studies have suggested that serum protein patterns may be able to distinguish cancer patients from control subjects. We used matrix-assisted laser desorption and ionization (MALDI) mass spectroscopy to obtain serum protein patterns from patients with head and neck cancer (n = 99) or lung cancer (n = 92) and from control subjects (n = 143) at risk for the development of these cancers. From the mass spectra, we predicted the cancer status of patients using a simple classification procedure based on a t test feature selection and linear discriminant analysis (LDA). We cross-validated the data with 200 random data simulations to establish a range of the LDA tuning parameter, which was used to construct receiver operating characteristic (ROC) curves. Average total protein levels were higher in case patients than in control subjects, although the differences were not statistically significant. Ten individual m/z peaks, from 5 to 111 kd, appeared frequently in head and neck cancer patients but not in control subjects. Using the 45 top predictors, selected by spectral mass and LDA, we observed that ROC curves differed from those expected under the null hypothesis, suggesting that spectral profiles from the sera of patients with head and neck cancer statistically significantly differed from the sera of control subjects. The model developed on head and neck cancer patients could also be used to identify patients with lung cancer. The pattern of protein spectra in total serum reliably distinguished cancer case patients from control subjects. Incorporation of MALDI assays into prospective longitudinal trials to assess the true predictive values of protein spectra in cancer detection is needed.

Evaluation of IgG4 response in ascariasis by ELISA for serodiagnosis.

The excretory/secretory (ES) antigen(s) of Ascaris lumbricoides was fractionated into 10 fractions by gel chromatography on a Suparose 12 column in FPLC. Of these, the third fraction (Al III), showing binding activity with both IgE and IgG antibodies of A. lumbricoides infected patients' sera, was further resolved into 2 fractions (Al IIIa and Al IIIb) on passage through a Mono Q column. Al IIIb was found to be the most potent antigen due to its high binding affinity with IgE and IgG antibodies of Ascaris infected patients as evidenced by ELISA inhibition. Although a two to five-fold increase of serum IgE level was observed in all helminthic parasite infected patients studied compared to control subjects, Al IIIb specific IgE was detected in sera of all Ascaris infected and only 40% of hookworm infected patients. When Al IIIb was tested by ELISA with sera of control subjects, Ascaris, hookworm, Strongyloides and Trichuris infected patients, strong binding was observed with the IgE and also IgG of all the Ascaris infected patients; however, it cross-reacted with IgG in 50% of hookworm, 28.6% of Trichuris trichura and 22.2% of Strongyloides infected patients' sera; but with IgE only in 40% of hookworm infected patients' sera. Further study showed specific detection of IgG4 in all the serum samples of 65 Ascaris infected patients when Al IIIb antigen was allowed to react with different subclasses of IgG by ELISA, giving a sensitivity of 100%. Reactivities of Al IIIb with IgG1 and IgG3 were only 47.6 and 11.8% respectively and there was no reactivity with IgG2 subclass. No IgG4 reactivity against Al IIIb was observed in the sera of hookworm, Trichuris or Strongyloides infected patients and was similar to that observed with control subjects showing the 100% specificity of the test system. This study may therefore be regarded as a novel technique for serodiagnosis of ascariasis by measuring Ascaris specific IgG4.

High levels of sTNFR p75 and TNF alpha in dengue-infected patients.

Soluble forms of the two molecular species of the cell surface TNF receptors (sTNFR p55 and sTNFR p75) can reduce the activity of TNF alpha but they may also enhance its function by stabilizing the active TNF alpha oligomer. Considering the pathophysiological importance of sTNFR p75 for the regulation of the bioavailability of TNF alpha in the body, we determined the serum levels of sTNFR p75 and TNF alpha in 45 children and 28 adults with laboratory-confirmed dengue infection by using immunoassays. The serum samples were obtained from day 1 to day 15 after the onset of the disease during the 1989-1990 outbreak of dengue-3 in Tahiti, French Polynesia. The patients were clinically classified as having dengue hemorrhagic fever (DHF) and graded according to the criteria of the World Health Organization (WHO) into four grades from less severe (grade I) to severe (grade IV). The sera of both children and adult patients of all severity grades contained higher levels of sTNFR p75 than the sera of control subjects. Although high levels of TNF alpha were also detected in children and adults among grade I, II, III and IV patients, we found no correlation between sTNFR p75 and TNF alpha. We observed in adults a moderate elevation of sTNFR p75 and TNF alpha in sera compared with that observed in children. The raised levels of immunoreactive sTNFR p75 and TNF alpha in all clinical groups of dengue-infected patients strongly indicate activation of the TNF alpha system during dengue infection. The balance between sTNFR p75 and TNF alpha may be altered in dengue infection. Further investigations are needed to understand the role of sTNFR p75 and TNF alpha in the pathogenesis of DHF and to improve the management of dengue infection.

Cockroach-allergen study: Allergen patterns of three common cockroach species probed by allergic sera collected in two cities

Antigens/allergens of three common cockroach extracts, crude whole body extract of the American cockroach (CRa-A), crude whole body extract of the German cockroach (CRa-G), and crude whole body extract of the Oriental cockroach (CRa-O), were studied with crossed immunoelectrophoresis, crossed radioimmunoelectrophoresis, and Western blot analysis. Sera of cockroach-allergic patients with asthma, 10 from Chicago, Ill. (C group) and six patients from Lexington, Ky. (L group), were used; results were then compared with sera of control subjects with asthma. Qualitative differences in protein bands were noted among CRa-A, CRa-G, and CRa-O by crossed immunoelectrophoresis and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Allergen bands on Western blot were analyzed for distribution by molecular weight (MW) with relative intensity scores. Results were compared by species and by geography. Two to 12 allergenic bands of variable MW (14 kd to >116 kd) were identified by 13 of 16 individual sera from cockroach-allergic patients from all three extracts. CRa-A demonstrated 55 bands with an intensity score of 125; CRa-G, 58 bands with an intensity score of 100; and CRa-O, 51 bands with an intensity score of 108. Allergenic bands of CRa-A were identified by six sera of the C group and one sera of the L group, whereas bands of both CRa-G and CRa-O were noted by nine sera of the C group and four sera of the L group. All three species had an allergen band in MW range of 40 to 45 kd that reacted to most sera from cockroach-allergic patients with asthma.

Evaluation of the fructosamine test in obesity: consequences for the assessment of past glycemic control in diabetes

The effect of obesity in diabetic and nondiabetic states on serum fructosamine levels, as measured by the nitro blue tetrazolium reduction method, was investigated. In 26 nondiabetic obese subjects, the mean (SD) fructosamine (1.78 +/- 0.16 mmol/L) and protein corrected fructosamine concentrations (25.7 +/- 2.5 mumol/g) were significantly lower than in nondiabetic lean control subjects (2.06 +/- 0.18 mmol/L and 30.5 +/- 2.5 mumol/g, respectively; p less than 0.01). Hemoglobin A1C, blood glucose and serum protein concentrations were normal in obese subjects. Interference from hypertriglyceridemia, hemolysis, or drugs was excluded. In diabetic subjects, fructosamine correlated with hemoglobin A1C, but the least-squares regression lines were different in 16 nonobese and in 19 obese patients, so that for the same hemoglobin A1C value, fructosamine level was 16% lower in obese compared to nonobese diabetic subjects. In vitro studies showed a significant decrease in 14C-glucose incorporation in serum proteins of obese nondiabetic subjects compared to control subjects. Similarly, the rate of formation of fructosamine in sera of obese nondiabetic subjects incubated with 12 mmol/L and 30 mmol/L glucose concentrations was slower than in sera of control subjects. In conclusion, fructosamine is underestimated in obesity, both in diabetic and nondiabetic patients, and its validity as an index of glycemic control may be impaired in obese subjects. This decrease is due to an alteration in the glycation process itself.

Axonal neuropathy with monoclonal IgG kappa that binds to a neurofilament protein

We report a 74-year-old woman with a slowly progressive sensory motor axonal neuropathy and a monoclonal IgG-kappa that bound to a 68-kd axonal protein identified as the low molecular weight neurofilament protein. The sera of control subjects and disease controls did not bind to neurofilament protein.

Decreased serum β- D-mannosidase activity in diabetic patients, in comparison with other glycosidases

A study of four lysosomal glycosidases' activities was carried out on sera from 64 diabetic patients, which revealed important variations in comparison with the activities observed in sera of control subjects. Depending on the type of diabetes mellitus (I, insulin-dependent, or II, non-insulin-dependent), three activities were more or less increased: α- L-fucosidase, α- D-mannosidase, and N- acetyl-β- D - glucosaminidase , in agreement with previously published results. Against that, the β- D-mannosidase activity shows a highly significant decrease in sera from either diabetic type. Up to now, no suitable explanation has been found for these variations occurring in an unusual direction.

The enzyme linked immunosorbent assay in the immunodiagnosis of human malaria

The sera of 20 malaria ( Plasmodium vivax) positive patients were examined for the presence of antibodies by enzyme linked immunosorbent assay (ELISA) using heterologous P. knowlesi antigen collected from parasitized erythrocytes from a Rhesus monkey ( Macaca mulatta). All the samples from malaria patients gave a strong ELISA test. The test was negative or weak with the sera of control subjects.

Serum antibody findings in trichinosis (author's transl)

Serum samples of 66 persons who had eaten meat from a wild boar were tested two to three times within 11 months for antibodies against extracts from Trichinella larvae. Watery extract of Trichinella spiralis larvae served as antigen. Complement-fixation was of little use because of its lack of sensitivity. Indirect haemagglutination and the ELISA test (enzyme-linked immunosorbent assay) gave uncertain results only in individual cases. There was a relationship between the clinical picture and the mean antibody concentration, clearer in the ELISA test than the indirect haemagglutination one. A fall in antibody titre was observed within the period of observation in only one third of patients. Results with sera of control subjects were of only limited use in relation to the specificity of the demonstrated antibodies.

Development of the hyaluronidase activity assay as a cancer screening test

1. Hyaluronidase activity, determined from the concentration of N-acetylglucosamine end groups of split products of hyaluronic acid, was on the average lower in the sera of patients with advanced carcinomas and hemoblastoses than in the sera of control subjects. Normal activities were observed in patients with metastatic sarcomas. 2. The greatest decrease in localized carcinomas was found in patients with disease of the genital, urinary and colorectal organs. Attempts to demonstrate serum hyaluronidase inhibitors in cancerous sera by the colorimetric assay were unsuccessful.