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J Kleine-Tebbe

Publications and source records attributed to J Kleine-Tebbe.

At least 37 records · Page 2Linked to original sources
PubMed1996

Comparison between two automated systems to determine specific IgE: CAP and ELItest.

BACKGROUND AND OBJECTIVE: The ELItest is a newly developed system to measure specific IgE based on allergen bound to paper rings and an alkaline phosphatase conjugated second antibody detection system. It was compared to the CAP system, a method based on allergen conjugated to an encapsulated cellulose polymer and a beta-galactosidase conjugated fluorescence detection system. METHODS: Sera of 300 patients with positive history and positive skin-prick tests to common allergens (birch, timothy-grass, cat dander, dermatophagoides ptronyssinus, wasp venom) and 30 negative controls were tested in both systems. Serial dilutions of high titre sera were measured; inter- and intraassay coefficients of variation (cv) were determined. RESULTS: The CAP system proved to be more sensitive (92.3%) compared to ELItest (84%) but marginally less specific (94.7% for CAP versus 96.7% for ELItest). Intraassay cv were slightly lower in the ELItest (7.2% CAP versus 6.4% ELItest), whereas the interassay cv was roughly twice as high for ELItest (20.1%) than for the CAP system (11.4%). Linearity over an 8-fold dilution was good in both tests (r2 0.979 ELItest versus 0.996 CAP), although ELItest levelled off at higher allergen concentrations. Similarly, correlation analysis between both systems revealed that ELItest consistently measured lower values, especially at higher concentrations of specific IgE. The slope of the linear regression line of a log/log plot of measured IgE concentrations was significantly lower than 1 in birch, cat and wasp; the y-intersect was significantly lower than 0 in all analysed allergens. CONCLUSION: These results suggest that the ELItest system for the measurement of specific IgE is not quite as reproducible and sensitive as the CAP system but slightly more specific, and that higher concentrations of specific IgE are measured lower in the ELItest. One potential reason might be that the amount of allergen bound to a paper ring might be smaller than that bound to a cellulose polymer, but further experiments are necessary to prove this hypothesis.

Adolescent↗
PubMed1995

IL-13 expression at the sites of allergen challenge in patients with asthma.

Atopic asthma is characterized by inflammatory responses of the airway and is associated with up-regulation of Th2 cytokines, notably IL-4 and IL-5. A recently described human cytokine, IL-13, is a potent in vitro modulator of various cell types, including monocytes, B cells, and endothelial cells. Similar to IL-4, it is also involved in the induction of IgE synthesis. However, the in vivo expression and function of IL-13 and its relation to disease remain to be defined. Using a segmental allergen challenge model, we have examined the in vivo expression of IL-13 in the bronchoalveolar lavage (BAL) cells of atopic patients. We found a significant enhancement of both IL-13 transcripts and secreted proteins in the allergen-challenged BAL compared with the saline-challenged control sites of asthmatic and rhinitic patients. In contrast, the expression of IL-13 transcripts was not detected in the BAL of two normal subjects challenged with the same dose of ragweed allergen. The cellular source of IL-13 mRNA was identified in the mononuclear cell fraction of the allergen-challenged BAL. The allergen-induced quantitative differences in the level of transcripts were confirmed by competitive PCR assays. These results suggest that the significant increase in IL-13 in the allergen-challenged BAL is primarily from the mononuclear cells and is involved in the regulation of allergen-induced late phase inflammatory responses.

Adult↗
PubMed1995

Purification of immunoglobulin E (IgE) antibodies from sera with high IgE titers.

A three stage method for the ultrapurification of polyclonal IgE from human serum is reported using anion exchange chromatography followed by monoclonal antibody based positive and negative affinity chromatography. Following dialysis of 25-100 ml of serum (2.3-14 micrograms IgE/ml, n = 4) against 0.05 M Tris pH 8, each specimen was subjected to diethylaminoethyl (DEAE)-cellulose chromatography (serum/matrix = 1/4). IgE was eluted with 0.05 M Tris, 0.05 M NaCl pH 8, yielding an IgE recovery of 61-93%, with removal of approximately 90% of other serum proteins and an IgE purity ([IgE]/[Igs]) of 0.1-1.1%. After adjusting to 0.1 M NaCl and concentrating approximately 30-fold, the eluted IgE was further purified by affinity chromatography using a panel of IUIS/WHO-documented mouse monoclonal anti-human immunoglobulin antibodies (alpha hIg-MAbs). First, the IgE-enriched DEAE-cellulose chromatography fraction was incubated in a batch mode with two alpha hIgE-Fc MAbs (HP6029, HP6061) coupled to CNBr-Sepharose, CL-4B. IgE was eluted with 0.05 M glycine pH 2.8 and immediately neutralized. The IgE recovery was 32-52% and IgE purity was 72-97%. Silver-stained SDS-PAGE and noncompetitive solid-phase two-site immunoenzymetric assays for total human IgA, IgE, IgG and IgM indicated that IgA, IgG and IgM were the only contaminants. Next, the IgE was concentrated 10-30-fold in the presence of 0.1% HSA. One IgE specimen was ultrapurified in a batch mode by negative selection chromatography using three pairs of alpha hIg-MAbs (alpha hIgA: HP6111 + HP6123; alpha hIgG: HP6017 + HP6046; alpha hIgM: HP6081 + HP6083) coupled to CNBr-Sepharose, CL-4B. IgE purity increased from 91% to > 99.9% with approximately 70% recovery of IgE for this step. The ultrapurified IgE antibody was shown to be functionally reactive for allergen and Fc epsilon RI receptors on human basophils. We conclude that alpha hIg-MAbs are powerful tools to facilitate the affinity purification of functionally active human IgE from serum; however, when the analyte is present in low concentration, a carrier protein needs to be added to minimize non-specific loss of the material during this process.

Antibodies, Anti-Idiotypic↗
PubMed1994

Inhibition of IgE- and non-IgE-mediated histamine release from human basophil leukocytes in vitro by a histamine H1-antagonist, desethoxycarbonyl-loratadine.

Loratadine, a new nonsedating histamine H1-antagonist, has been shown to inhibit immunologic release of inflammatory mediators in addition to its H1-receptor blocking properties. After oral administration, the agent is metabolized primarily to desethoxycarbonyl-loratadine (DCL). The basic piperidine, DCL, is readily soluble in water, whereas the nonbasic urethane, loratadine, is insufficiently soluble in water for some in vitro investigations. Therefore we used the metabolite, DCL, to study its influence on in vitro leukocyte histamine release (LHR) in 24 allergic and 22 nonallergic subjects. IgE-mediated and calcium ionophore A23187-induced LHR were inhibited by DCL in a dose-dependent fashion (values of drug concentration to induce 30% inhibition after stimulation with inhalant antigen, anti-IgE, concanavalin A, and calcium ionophore A23187 were 6, 8, 5, and 11 mumol/L, respectively). Higher concentrations of DCL caused mediator release in all subjects (n = 45, 30 mumol/L DC: 11% +/- 2% LHR, 100 mumol/L DCL: 35% +/- 1% LHR), abolishing any inhibitory effect of the drug. Rapid onset of inhibition by 10 mumol/L DCL was found in kinetic studies (n = 10). The inhibition of anti-IgE-induced histamine secretion was synergistically increased by simultaneous preincubation of DCL with the potent histamine H2-agonist, FRA-19. Additional data indicate that the inhibition of LHR by DCL might involve biochemical events that occur after cellular Ca++ influx because LHR induced by N-formyl-methionyl-leucyl-phenylalanine or the phorbol ester, 12-O-tetradecanoyl phorbol-12-acetate, was not significantly affected by DCL.

Basophils↗
PubMed1994

Measurement of allergen-specific IgD and correlation with allergen-specific IgE.

A new method for the measurement of allergen-specific IgD (as-IgD) was developed by modifying the ImmunoCAP assay (Pharmacia), and amplification of the signal with a goat anti-human/rabbit anti-goat detection system. The assay was sensitive enough to measure as-IgD in serum samples. The specificity of the assay was examined using inhibition tests with excess corresponding and non-corresponding allergens. For the different allergens inhibition rates between 56% (house dust mite) and 88% (cat) could be achieved. Non-corresponding allergens did not inhibit the as-IgD binding. Total IgE and allergen-specific IgE (as-IgE) was measured using the ImmunoCAP system. Total IgD was measured using a sandwich ELISA. As-IgD was measured in serum samples from 51 atopic and 23 non-atopic subjects, and the correlation with as-IgE was examined. As-IgD was detected in both atopics and non-atopics but at higher levels in atopics. As-IgD against birch pollen and timothy pollen allergen was found to be increased in atopics with IgE directed against these allergens compared to atopics without IgE against these allergens (P < 0.02 and P < 0.03). As-IgD against birch pollen allergen was higher in atopics with IgE specific to this allergen than in non-atopics (P < 0.02). In contrast to total IgE and total IgD, significant correlations were observed between as-IgD and as-IgE against timothy pollen (r = 0.34, P < 0.04), birch pollen (r = 0.38, P < 0.05) and cat dander allergen (r = 0.52, P < 0.01). The observed correlations between as-IgD and IgE suggest that IgD and IgE may be similarly regulated, and thus the measurement of as-IgD may give further insight into the regulation of IgE.

Adolescent↗
PubMed1993

Measurement of membrane bound IgD and IgM on B lymphocytes.

An assay system for the determination of the membrane bound IgD (mIgD) and IgM (mIgM) on B lymphocytes was developed by the combination of two new ELISA methods with the results of flow cytometry after labeling with specific antibodies. The mIgD and mIgM of B lymphocytes were prepared by incubating mononuclear cells (MNCs) in Tween 20 containing buffer and repeated freeze/thaw cycles. Optimal results were achieved with 0.2-0.4% Tween 20 and two freezing cycles. With biotin-streptavidin amplification the sensitivity of the ELISA was 30 microU/ml for IgD and 0.5 ng/ml for IgM. In healthy persons 3.5 +/- 0.5 mU mIgD were detected on 10(6) IgD+ cells and 57.1 +/- 5.9 ng mIgM on 10(6) IgM+ cells. The mIgD/mIgM ratio was 0.065 +/- 0.005 mU/ng. The developed ELISA systems utilize only commercially available reagents and therefore provide a convenient reproducible tool for determining membrane bound IgD and IgM on B lymphocytes.

Adult↗
PubMed1993

Predominance of the major allergen (Alt a I) in Alternaria sensitized patients.

Thirty-nine patients sensitized to Alternaria were evaluated using titrated skin-prick test (SPT), histamine release studies (HR), inhibition of RAST and immunoblotting studies. To determine the relevance of the major allergen, Alt a I, specific rabbit antibodies against Alt a I and Alt a B were used. The antibodies were preincubated at different concentrations: (i) with the Alternaria allergen dose required for maximum response in the HR assay (10 BU/ml) and (ii) with the Alternaria antigen coupled to RAST paper discs (1000 BU/disc). Dose dependent inhibition of histamine release (n = 30, mean = 80% +/- 4%, IC30 = 0.69 micrograms/ml) and of RAST (n = 7, IC30 = 4.4 micrograms/ml) was found in all patients sensitized to Alternaria as indicated by allergen induced HR. The greater the response to Alternaria in HR, the higher the antibody concentrations necessary for inhibition (P < 0.05). Immunoblot experiments (n = 25) using SDS-PAGE showed in all cases IgE- and IgG binding at approximately 28 kD, which is the size reported for the major allergen, Alt a I. In two cases, slight IgE binding at 45 and 66 kD was also found, while in two other patients, only IgE binding at 66 kD was seen. Our findings emphasize the major importance of Alt a I in patients sensitized to Alternaria.

Adolescent↗
PubMed1993

Comparison of fiberglass-based histamine assay with a conventional automated fluorometric histamine assay, case history, skin prick test, and specific serum IgE in patients with milk and egg allergic reactions.

A microfiberglass-based histamine assay (HRM) was compared with an automated fluorometric histamine assay (HRA). Twenty-four subjects with and 24 without a case history (CH) of milk and/or egg allergy were tested by HRM and HRA, skin prick test (SPT), and specific serum IgE (RAST). Six different concentrations of milk, egg, and anti-IgE were used to stimulate washed leukocytes (250 microliters for HRA) and whole blood samples (25 microliters for HRM) in parallel. When we compared scores representing basophil sensitivity, correlation coefficients (rs) were positive (r(anti-IgE) = 0.88, r(egg) = 0.95, r(milk) = 0.88, P < 0.001), but no significant correlations were found after exclusion of the negatives in both tests. In some individual dose-response curves, the scores obtained by HRM were shifted to higher allergen and anti-IgE concentrations. A high degree of concordance was found in positive and negative responses between the two tests: anti-IgE 91%, egg 92%, milk 86%. Finally, we found a good concordance between, on one side, HRM and, on the other, CH, SPT, and RAST (HRM vs. CH/SPT/RAST): egg 92/82/82%; milk 89/74/67%. We conclude that HRM is in good qualitative, but poor quantitative, agreement with the autoanalyzer-based fluorometric histamine assay.

Adolescent↗
PubMed1993

Role of the major allergen (Fel d I) in patients sensitized to cat allergens.

Forty patients with a positive case history and specific serum IgE to cat allergens were evaluated using titrated skin prick test (SPT), histamine release studies (HR) and immunoblotting studies. To determine the relevance of the major allergen, a monospecific antibody to Fel d I was preincubated in different concentrations with the cat allergen dose required for maximum response in the HR assay (10 SQ/ml). Dose-dependent inhibition of histamine release (n = 30, x = 66% +/- 5% IC30 = 0.36 micrograms/ml) was found in all patients sensitized to cat allergens as shown by antigen-induced HR. The greater the HR response to the cat allergen extract the higher were the antibody concentrations necessary for inhibition (p < 0.05). Immunoblot experiments (n = 22) using SDS-PAGE demonstrated five different locations of IgE binding in 15 sera: 18, 35, 46, 54 and 66 kD. IgE binding at 18 kD was found in 14 patients. Anti-Fel d I possessed two strong bands at about 18 and 35 kD which are representative of the major allergen Fel d I. Our findings emphasize the major role of Fel d I in patients sensitized to cat allergens.

Adolescent↗
PubMed1992

Modulation of IgE-mediated histamine release from human leukocytes by a new class of histamine H2-agonists.

A new class of phenyl (pyridylalcyl) guanidines, acting as potent histamine H2-agonists, inhibits IgE-mediated human basophil histamine release in a nanomolar range. IC30-level of three substitutes of this group (arpromidine, BUA-75, and FRA-19) were found to be 0.02, 0.015 and 0.008 microM. The inhibition appeared with a fast onset (plateau after 10 min. preincubation) and claimed its maximum (60 +/- 2.9%, 63 +/- 1.8%, and 61 +/- 3.1%, n = 7) with 10 microM of the compounds. H2-mediated inhibition was totally blocked by 10 microM famotidine, a potent histamine H2-antagonist. The amount of anti-IgE or antigen for the initiation of the immunological release influenced the strength of inhibition of H2-agonist FRA-19 (p less than 0.05). Combined preincubation of FRA-19 with zardaverine, a cAMP-specific phosphodiesterase III/IV inhibitor, produced a synergistical inhibitory effect of leukocyte histamine release, which might explained by their different sites of action on intracellular cAMP levels. The capability of histamine to inhibit its own release is mediated by H2-receptors exclusively. New, potent H2-receptor stimulating compounds with positive inotropic effects possess additional potent anti-allergic properties.

Basophils↗
PubMed1992

Comparison between MAGIC LITE- and CAP-system: two automated specific IgE antibody assays.

The MAGIC LITE system, a newly developed immunochemiluminometric assay for specific and total IgE antibody using paramagnetic particles coupled with standardized allergens as solid phase, was compared to the CAP system, a recently introduced immunoassay based on a cellulose polymer encased in a capsule. A total of 357 serum samples of patients with suspected inhalant allergies and a positive skin prick test (SPT) to common allergens (birch, timothy-grass, mugwort, cat dander, Dermatophagoides pteronyssinus, Alternaria) were investigated. Fifty SPT negative subjects served as controls (total number of tests in each assay = 1600). Both assays were highly precise (overall intra-assay and inter-assay coefficients of variation were 2.9% and 4.5% in MAGIC LITE, 4.7% and 5.5% in CAP) and showed excellent linearity (mean r2 of eightfold log2 serum dilutions were 99.7% and 99.3% in MAGIC LITE and CAP). Good correlations were found between the absolute specific IgE antibody values detected by both methods (correlation coefficient r: birth 0.86, grass 0.93, mugwort 0.96, cat 0.91, D. pteronyssinus 0.73, Alternaria 0.90). Excellent specificity (greater than or equal to 98%) occurred in both assays and with all allergens, and sensitivity was related to the allergen (MAGIC LITE/CAP): birch 91%/89%, grass 83%/90%, mugwort 50%/69%, cat 83%/83%, D. pteronyssinus 72%/78%, Alternaria 75%/81%. Our results indicate that both in vitro tests are useful tools for the detection of specific IgE antibody.

Allergens↗
PubMed1992

Basophil histamine release in patients with birch pollen hypersensitivity with and without allergic symptoms to fruits.

Histamine release (HR) studies were performed in 40 birch pollen-allergic patients (positive case history, positive SPT, positive birch pollen-specific serum IgE: RAST > or = 3) with (n = 20, A) and without (n = 20, B) fruit hypersensitivity, and 10 nonatopic volunteers (C). Several fruit allergens were used and characterized by protein determination and immunoblot techniques. Dose-dependent HR (apple peel = apple pulp > peach = cherry) was demonstrated in both allergic groups, but to a higher extent in patients with fruit allergy (P < 0.01). Increased basophil sensitivity to birch pollen was found in the group with fruit allergy (P < 0.001). Strong correlations between the mediator response induced by several fruits indicate common allergens within the extracts. We conclude that fruit-related symptoms require not only high specific serum IgE, but a strong cellular sensitization to birch pollen allergens together with an increased cellular reactivity to fruit allergens.

Adult↗
PubMed1992

Inhibition of IgE-mediated histamine release from human peripheral leukocytes by selective phosphodiesterase inhibitors.

Several phosphodiesterase (PDE) inhibitors with the capacity to inhibit the PDE IV isoenzyme produce dose-dependent inhibition of IgE-mediated histamine release (HR) from human peripheral leukocytes in vitro. Inhibition reached a maximum after 20 min of preincubation (IC30: 6-30 microM, IC50: 30-80 microM). Motapizone--a potent inhibitor of the isoenzyme PDE III--was much less effective, thus giving indirect evidence that PDE IV plays a predominant role in the control of cAMP cleavage in human basophils. The inhibiting effect of PDE-III/IV-selective compounds on IgE-mediated HR did not exceed the action of PDE-IV-selective inhibitors. The inhibition of anti-IgE-induced HR by zardaverine (a PDE-III/IV-inhibiting compound) was synergistically enhanced in the combined presence of forskolin or the recently synthesized histamine H2-agonist FRA 19).

Basophils↗
PubMed1992

IgE-mediated inhalant allergy in inhabitants of a building infested by the rice weevil (Sitophilus oryzae).

The rice weevil (Sitophilus oryzae) is known as a stored product pest. Occupants living in a factory used previously as a granary and known to be contaminated by the rice weevil developed rhinitis and asthma. To study the role of the potential insect allergens, extracts were prepared from whole body and a grain dust of the rice and grain weevil (Sitophilus granarius). The extracts were coupled to cyanogen bromide-activated paper disks. Skin prick tests (SPT) were performed using common inhalant allergens as well as the insect allergen preparations. In cases of a positive SPT, specific serum IgE to the insect products was measured. Histamine release studies with peripheral leukocytes (HR) and studies using immunoblot techniques were performed. Fifteen of 39 subjects living in the infested rooms demonstrated positive SPT to the extracts of insect origin. Histamine equivalent wheal size induced by the whole body extract of the rice weevil required 3-160 micrograms protein/mL. Five of 15 subjects (with positive SPTs) had elevated specific IgE-levels to the insect material. RAST-inhibition studies indicated cross allergenicity between rice and grain weevils, and also between the frass of both beetles. Five of 15 subjects were positive in the HR assay (Ag30, allergen concentration for 30% HR for the whole body extract: range, 10(-3) to 400 micrograms protein/mL, maximum release: 26% to 89%). IgE binding was detected after SDS-PAGE by immunoblot techniques at different locations: 35-38, 54, 67, 70, and > 94 kD.(ABSTRACT TRUNCATED AT 250 WORDS)

Administration, Inhalation↗
PubMed1991

The possible role of substance P in the allergic reaction, based on two different provocation models.

It was shown in two different provocation models (nasal and bronchial provocation) that substance P (SP) may play an important role in the neurogenic inflammatory response in upper and lower airway disease. (1) Pretreatment with SP augments the antigen challenge response of the nasal mucosa. (2) The baseline bronchoalveolar lavage (BAL) concentrations of SP are elevated 8-fold in allergies (pollen asthma) as compared with normals, even outside of season. (3) The SP concentration in BAL increases significantly (p less than 0.05) after bronchial allergen provocation. These findings support a previous hypothesis of an abnormally elevated activity of nonadrenergic-noncholinergic excitatory nerves and are in accordance with the results of a decreased activity of neutral endopeptidase exaggerating neurogenic inflammatory responses in the airways, including bronchomotor tone hyperresponsiveness.

Adult↗
PubMed1991

IgE-mediated inhalant allergy against human corticotropin-releasing hormone.

A 25-year-old female developed IgE-mediated sensitization against human recombinant corticotropin-releasing hormone (CRH) with symptoms of allergic rhinoconjunctivitis and bronchial asthma. The occupational allergy was proved by positive skin prick test, bronchial provocation, dose-dependent histamine release, RAST measurements with CRH allergen (RAST class 3) and RAST inhibition. Using the immunoblot technique, a single allergen band with a molecular weight of less than 14.4 kD in the range between the isoelectric point 5.2 and 5.7 was detected for the CRH extract. Since no endocrinological and behavioral disorders were found, increased CRH-specific IgE was not able to influence the regulatory control of this neuropeptide. After 18 months of avoiding the occupational CRH exposure allergen-specific histamine release and RAST were negative.

Adult↗
PubMed1990

Influence of histamine H3-antagonists on human leukocytes.

To determine the role of the histamine H3-receptor on basophils, different specific H3-antagonists were investigated. Incubation of washed leukocytes with N alpha-acylated histamine-derivatives (N alpha-ahd) induced elevated histamine levels. This process turned out to be dependent on dose, time and temperature, but independent of Ca2+ and Mg2+ ions. IgE-mediated histamine release was not modulated. [3H]-L-histidine was not decarboxylated into [3H]-histamine in spite of the observed histamine increase. Highly purified basophils did not show any histamine elevation but purified neutrophils and eosinophils were found to have increased histamine levels even after disintegration and subsequent incubation with N alpha-ahd. It seems that the increased histamine levels result from the cleavage of the applied histamine amides. Other potent H3-antagonists (e.g. thioperamide) neither produced increased histamine levels nor influenced IgE-mediated release from basophil leukocytes. The existence of H3-receptors on human basophils therefore seems unlikely.

Basophils↗