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M Aalbers

Publications and source records attributed to M Aalbers.

16 recordsLinked to original sources
PubMed2004

A multi-laboratory evaluation of a common in vitro pepsin digestion assay protocol used in assessing the safety of novel proteins.

Rationale. Evaluation of the potential allergenicity of proteins derived from genetically modified foods has involved a weight of evidence approach that incorporates an evaluation of protein digestibility in pepsin. Currently, there is no standardized protocol to assess the digestibility of proteins using simulated gastric fluid. Potential variations in assay parameters include: pH, pepsin purity, pepsin to target protein ratio, target protein purity, and method of detection. The objective was to assess the digestibility of a common set of proteins in nine independent laboratories to determine the reproducibility of the assay when performed using a common protocol. Methods. A single lot of each test protein and pepsin was obtained and distributed to each laboratory. The test proteins consisted of Ara h 2 (a peanut conglutin-like protein), beta-lactoglobulin, bovine serum albumin, concanavalin A, horseradish peroxidase, ovalbumin, ovomucoid, phosphinothricin acetyltransferase, ribulose diphosphate carboxylase, and soybean trypsin inhibitor. A ratio of 10U of pepsin activity/microg test protein was selected for all tests (3:1 pepsin to protein, w:w). Digestions were performed at pH 1.2 and 2.0, with sampling at 0.5, 2, 5, 10, 20, 30, and 60min. Protein digestibility was assessed from stained gels following SDS-PAGE of digestion samples and controls. Results. Results were relatively consistent across laboratories for the full-length proteins. The identification of proteolytic fragments was less consistent, being affected by different fixation and staining methods. Overall, assay pH did not influence the time to disappearance of the full-length protein or protein fragments, however, results across laboratories were more consistent at pH 1.2 (91% agreement) than pH 2.0 (77%). Conclusions. These data demonstrate that this common protocol for evaluating the in vitro digestibility of proteins is reproducible and yields consistent results when performed using the same proteins at different laboratories.

Clinical Laboratory Techniques↗
PubMed2000

Reactivity to IgE-dependent histamine-releasing factor is due to monomeric IgE.

BACKGROUND: IgE-dependent histamine-releasing factor (HRF) can distinguish between IgE+ and IgE-. In contrast to IgE-, IgE+ sensitizes basophils to release histamine in response to HRF. But we do not know what particular feature distinguishes IgE+ from IgE-. The objective was to investigate the hypothesis that IgE+ is polymeric IgE. METHODS: IgE+ plasma was separated by size-exclusion chromatography. The basophil-sensitizing capacity of the fractions was analyzed in response to HRF produced by mononuclear cells. RESULTS: We showed that monomeric IgE sensitized basophils to release histamine in response to HRF and to house-dust mite, whereas no enhanced reactivity was found in the fractions containing polymeric IgE. CONCLUSIONS: HRF reacts with monomeric IgE, and not (exclusively) with polymeric IgE.

Antigens, Dermatophagoides↗
PubMed2000

A sensitive monoclonal antibody sandwich ELISA for the measurement of the major olive pollen allergen Ole e 1.

BACKGROUND: Olive pollen is a major cause of inhalant allergy in countries around the Mediterranean sea. The major allergen of olive pollen is Ole e 1. Measurement of the major allergen content of allergen products for diagnosis and therapy is becoming an essential element of standardization protocols. This study aimed at the development of a monoclonal antibody (mAb) sandwich ELISA for Ole e 1. METHODS: Balb/c mice immunized with Ole e 1 were used for the production of mAbs. Screening of mice and hybridomas was performed in a RIA with radiolabeled purified Ole e 1. Purified mAbs were used as catching and/or (biotinylated) detecting antibodies in sandwich ELISA. RESULTS: Four mAbs (IgG1kappa) directed to nonoverlapping epitopes on Ole e 1 were obtained: 1A12, 5C1, 10A12 and 3H8. Both 1A12 and 10A12 were successfully used for affinity purification of Ole e 1 from olive pollen extract. Two sandwich ELISAs were developed, with 1A12 and 10A12 as catching, and 5C1 and 3H8 as detecting antibodies, respectively. Both catching and detecting antibodies were used in similar concentrations, ranging from 60 to 100 ng/well. For both ELISAs, the sensitivity was approximately 1 ng/ml of Ole e 1. The measuring range was from 1 to 25 ng/ml. No significant differences were observed, when the performance of both ELISAs in standardization of olive pollen extracts was compared. CONCLUSIONS: Two sensitive sandwich ELISAs for the major olive pollen allergen Ole e 1 were developed. They will prove to be useful tools in allergen standardization protocols.

Allergens↗
PubMed1996

Histamine-releasing activity in supernatants of mononuclear cells: contribution of monocyte chemotactic protein-1 activity compared with IgE-dependent activity.

BACKGROUND: Supernatants of cultured human mononuclear cells contain factors that induce histamine release from basophils. Some of the histamine-releasing factors present are IgE-independent, but an IgE-dependent form has also been described. The IgE that does respond to the IgE-dependent histamine-releasing factor was defined as IgE+. OBJECTIVE: This study was conducted to analyze the contribution of chemokines, such as monocyte chemotactic protein-1 (IgE-independent), to improve the detection of IgE-dependent histamine-releasing factor. METHODS: Supernatants were prepared from mononuclear cells of three subjects. Monocyte chemotactic protein-1 content and histamine-releasing activity (HRA) were measured. Depletion of chemokine activity was carried out with heparin-Sepharose (Pharmacia, Uppasala, Sweden). RESULTS: Replacing IgE- on basophils with IgE+ serum decreased the correlation between monocyte chemotactic protein-1 and histamine release (rho = 0.80, n = 280 vs p = 0.12, n = 18; p for difference between p values; < 0.05). After depletion of chemokines in three supernatants derived from mononuclear cells, IgE-dependent HRA was still present. CONCLUSION: We conclude that in supernatants derived from mononuclear cells, the IgE-independent HRA masks the IgE-dependent HRA. The latter can be more clearly detected after depletion of chemokine activity with heparin-Sepharose.

Animals↗
PubMed1996

Reactivity to IgE-dependent histamine-releasing activity in asthma or rhinitis.

We investigated the relation between IgE reactive with histamine-releasing factor (HRF) and clinical status in patients with asthma or rhinitis. Sera were used to passively sensitize purified, lactic-acid treated basophils. IgE-independent HRF due to chemokines was removed from mononuclear cell supernatants with heparin-Sepharose. IgE-dependent HRF was determined by measuring the increase in histamine release between 1 min and 60 min, which was designated delta HRF. HRF-reactive IgE was demonstrable in nine of 18 patients with allergic asthma, three of 19 patients with nonallergic asthma, five of 17 patients with allergic rhinitis, and none of 19 control patients. The presence of HRF-reactive IgE was associated with: (1) IgE to inhalant allergens; 40% of radioallergosorbent test (RAST)-positive individuals versus 8% of RAST-negative individuals were positive (OR = 7.8, p < 0.005); (2) bronchial sensitivity to histamine in all asthmatic patients (geometric mean PC20: 1.50 versus 0.51 mg/ml; p < 0.005); and (3) bronchial sensitivity to histamine in allergic asthmatic patients (geometric mean PC20: 1.27 versus 0.37 mg/ml, p < 0.02). These findings support the hypothesis that IgE-dependent HRF might contribute to the chronic allergic reaction.

Adolescent↗
PubMed1995

Multiplicity of cross-reactive epitopes on Bet v I as detected with monoclonal antibodies and human IgE.

Six monoclonal antibodies against Bet v I, the major cross-reactive allergen of birch pollen (Betula verrucosa), were obtained. Four did not react with fruits, but two monoclonal antibodies (mAbs) (5H8 and 9C11) were reactive with apple and other fruits. These two cross-reactive antibodies reacted with identical or overlapping sites, but differed in their relative degree of cross-reactivity toward various fruits and hazelnut. Cross-reactive human IgE antibodies reacted with a nonoverlapping epitope, as indicated by results of a two-site radioimmunoassay (RIA) with the fruit-reactive mAb 9C11. By isoelectric focusing (IEF) in conjunction with immunoblotting, a maximum of seven isoforms could be distinguished. Depletion of birch-pollen extract for Bet v I with the most reactive mAb (7F7) removed approximately 95% of the IgE cross-reactivity between birch pollen and apple extract. The remaining 5% cross-reactive material was still capable of inhibiting the binding of IgE to apple allergen completely, and was reactive with mAbs 5H8 and 3C4. By means of IEF/immunoblot, it was shown that these mAbs recognize an isoform of Bet v I that is poorly, if at all, recognized by mAb 7F7. These results illustrate the heterogeneity of Bet v I, both with respect to the cross-reactive sites as well as to the backbone structure. This type of heterogeneity has possible implications for the use of monoclonal antibodies in allergen standardization.

Adult↗
PubMed1994

Variability of IgE-dependent histamine-releasing activity in supernatants of human mononuclear cells.

Histamine-releasing factors (HRF) that release mediators from human basophils by interacting with IgE have been identified from different cell sources, including lymphocytes, monocytes, thrombocytes and endothelial cells. These factors are studied in view of their potential importance as a stimulus in chronic inflammation. In this report we investigated the qualitative variability of the histamine-releasing activity in the supernatants of activated mononuclear cells. Purified human mononuclear cells of 8 donors were activated with streptokinase/streptodornase (SK/SD) and the supernatants (HRF-MN) were tested for histamine-releasing activity (HRA) in both allergic (RAST positive for inhalant allergens) and nonallergic individuals. Four of the eight HRF-MN supernatants were discriminating, i.e. showing no histamine-release response with nonallergic individuals, whereas four supernatants were not. Two of the HRF-MN supernatants that exhibited discriminating properties were studied in more detail. The response to HRF-MN was tested (1) in a direct bioassay on basophils of allergic (RAST positive for inhalant allergens) and nonallergic individuals and (2) in an indirect bioassay with 70% pure basophils of RAST-negative donors after passive sensitization with sera of allergic donors. An association was found between the response to HRF-MN and the RAST for inhalant allergens: none (0/12) of the RAST-negative but 15/22 of the RAST-positive individuals were HRF-MN responders. The IgE dependency of HRF-MN was shown e.g. by inhibition of passive sensitization by preincubating a responder serum with monoclonal antibody (moAb) anti-IgE MH25-1. Our results are in contrast with findings of other investigators who use pooled supernatants and demonstrated HRF-MN responsiveness with both allergic and nonallergic donors.(ABSTRACT TRUNCATED AT 250 WORDS)

Basophils↗
PubMed1992

Food intolerance in rheumatoid arthritis. II. Clinical and histological aspects.

Six patients with rheumatoid factor positive rheumatoid arthritis who had shown a marked symptomatic improvement during four weeks of hypoallergic, artificial diet were studied in greater detail. Placebo controlled rechallenges showed intolerance for specific foodstuffs in four patients. In three of these patients biopsies of both the synovial membrane and of the proximal small intestine were carried out before and during allergen free feeding. In two patients, both with raised serum IgE concentrations and specific IgE antibodies to certain foods, a marked reduction of mast cells in the synovial membrane and proximal small intestine was demonstrated. Although the number of food intolerant patients with RA remains limited and markers of allergic activity are scanty, our observations suggest an underlying immunoallergological mechanism.

Aged↗
PubMed1992

The potent IgG4-inducing antigen in banana is a mannose-binding lectin, BanLec-I.

IgG4 antibodies to banana were found to occur far more frequently than expected. The most important antigen involved proved to be a lectin, BanLec-I. Because of the lectin nature of the antigen, it was important to establish the antibody nature of the lectin-IgG4 interaction and to exclude an interaction between the sugar-binding site of the lectin and glycosidic chains on IgG4. Three arguments in support of immune binding are: (1) the binding of BanLec-I to IgG4 is mannoside resistant, whereas the binding to all other glycoproteins tested is mannoside inhibitable; (2) only a minor fraction of the IgG4 in serum and none of five IgG4 myelomas tested was bound, and (3) the lectin binds to the Fab fragment of the IgG4 molecule. A curious finding was that in the presence of high-molecular-weight glycoproteins the interaction between IgG4 and BanLec-I was enhanced by alpha-methyl mannoside. The probable explanation of this phenomenon is that complexes of the lectin with high-molecular-weight glycoproteins by sterical interference inhibit the interaction with human IgG4 antibodies (or with rabbit antibodies to the lectin). This inhibition is prevented in the presence of alpha-methyl mannoside. These results support the earlier suggestion that some lectins are particularly prone to induce an immune response upon oral feeding. This banana lectin might be a potentially useful carrier protein for oral antihapten immunization in humans.

Antigens↗
PubMed1991

Newly generated IgE antibodies to Dermatophagoides pteronyssinus in children are directed against components distinct from Der p I and Der p II.

The specificity of newly generated IgE antibodies (Abs) to the house dust mite, Dermatophagoides pteronyssinus, in longitudinal serum samples from 18 young children with an increased risk for IgE-mediated allergy was studied. The first IgE Ab response to house dust mite was detected early in life (mean age, 32 months; range, 11 to 60 months). For 83% of the children, more than half of the newly generated IgE Ab response to house dust mite was directed against components distinct from the major allergens, Der p I (Pl) and Der p II (DpX). These results suggest that the early IgE Ab response to house dust mite is induced by components distinct from the major allergens, Der p I and Der p II.

Allergens↗
PubMed1991

Relationship between IgG1 and IgG4 antibodies to foods and the development of IgE antibodies to inhalant allergens. I. Establishment of a scoring system for the overall food responsiveness and its application to 213 unselected children.

To obtain reference levels for subsequent investigations, we analysed the IgG1 and IgG4 antibody levels to common foods in the sera of 213 unselected children (age 3 months to 14 years). The children were clustered into five age groups and tested on a broad screening panel of common foods. We used the IgG1 and IgG4 RAST with Sepharose-coupled antigens: cows' milk, hens' egg white, banana, legumes (a mixture of soybean and peanut), grains (a mixture of wheat and rice), potato, orange and pork. In all age groups and all antigens, a considerable variability in the antibody response was found. As for some assays more than half of the sera were negative or borderline, statistics based on interval or ordinal scaling were considered inappropriate and we resorted to nominal classification. We decided to use, for each of the assays, the 75-percentile of the age group as a cut-off level. Each antibody titre was thus converted into positive (more than the 75-percentile of that age group) or negative; the number of positive tests was used as the score. This resulted in a sigma G1-score and a sigma G4 score (summed scores for IgG1 and IgG4 antibodies, respectively). The results of the present study indicate that children with a high response to one food tend to have elevated responses to other non-related foods, possibly explained by a defective mucosal barrier and/or a hyperactive immune system. This suggests that a high-food responder phenotype may exist.

Adolescent↗
PubMed1991

Relationship between IgG1 and IgG4 antibodies to foods and the development of IgE antibodies to inhalant allergens. II. Increased levels of IgG antibodies to foods in children who subsequently develop IgE antibodies to inhalant allergens.

In the present investigation we have tested the hypothesis that children with a high IgG antibody response to foods have an increased risk of developing IgE antibodies to inhalant allergens. Sera from 106 children with an increased risk of developing IgE-mediated allergy were analysed. During the follow-up, in 54 of these children IgE antibodies to inhalant allergens appeared. A positive/negative IgG1 and IgG4 anti-food score was determined as described previously: sera from age-clustered unselected children were tested for the levels of IgG1 and IgG4 antibodies to common foods. For each IgG RAST and each age group, the 75-percentile was chosen as cut-off value. Each antibody level was thus converted into a positive (higher than the 75-percentile of the age group) or negative value. The number of positive tests was used as the score. High-risk children with a high IgG1 anti-food score more often developed inhalant-specific IgE antibodies than high-risk children with low IgG1 titres: 50% of the children with a high IgG1 anti-food score developed IgE antibodies to grass pollen. Fifty per cent of the children with a high and 14% of the children with a low IgG1 anti-food score developed IgE antibodies to cat dander. For the prediction of the development of IgE anti-mite (house dust mite), the IgG4 anti-food scores appeared less useful than the IgG1 anti-food scores; 46% of the IgG4 high responders versus 22% of the IgG4 low responders acquired IgE anti-mite, whereas for IgG1 these percentages were 73 and 19, respectively.

Allergens↗
PubMed1989

Characterization with monoclonal and polyclonal antibodies of a new major allergen from grass pollen in the group I molecular weight range.

We have purified a 24/25 kd allergen from orchard grass pollen (Dactylis glomerata) that has an allergenic potency similar to that of the major group I allergen. We provisionally named this allergen grass 4B1 after the monoclonal antibody used for its identification and purification. This monoclonal antibody was obtained by immunizing mice with whole Lolium perenne-pollen extract and by screening the antibody producing hybrids for reactivity with Dactylis glomerata-pollen extract. Grass 4B1 is physicochemically separable from the group I allergen. Polyclonal rabbit antibodies to grass 4B1 do not react with group I allergen or vice versa. Ninety-five sera with IgE antibodies to grass pollen were tested for IgE antibodies to grass 4B1, and greater than 90% was positive in this test. The median response to grass 4B1 was 70% of that to Lol p I.

Allergens↗
PubMed1987

Cross-reactivity among birch pollen, vegetables and fruits as detected by IgE antibodies is due to at least three distinct cross-reactive structures.

Sera of patients suffering from birch pollinosis were studied in the radio-allergo-sorbent test (RAST) for the presence of IgE antibodies to various allergens of vegetable origin. The sera selected were positive in the RAST for both birch pollen and fruits. IgE antibodies directed against at least three different cross-reacting determinants in birch pollen were detected. In addition to periodate-susceptible cross-reacting determinants, which are found on a number of glycoproteins, two non-related periodate-resistant determinants were found in birch pollen, with molecular weights of 20 and 18 kD, respectively. The 20-kD component appears to be responsible for the co-occurrence of the binding of IgE to allergens of fresh fruits, whereas the 18-kD component appears to cause the cross-reactivity among grass pollen, potato and fruits.

Adult↗