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Nahrungsmittelallergie: Identifizierung und Charakterisierung von Erdnußallergenen mit Patientenseren und monoklonalen Antikörpern

Food allergy: Identification and characterization of peanut allergens with patients' sera and monoclonal antibodies

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Zusammenfassung

Zielsetzung dieser Arbeit ist eine verbesserte Diagnostik der Erdnußallergie. Um eine Standardisierung der Testsubstanzen für die in vivo- und in vitro-Diagnostik zu erreichen, wurden die Typ I Allergie assoziierten Einzelkomponenten der Erdnuß mit Hilfe von Patientenseren und monoklonalen Antikörpern identifiziert und charakterisiert. Zur Allergenidentifizierung wurden IEF-Immunoprint-, SDS-PAGE-Immunoblot- und 2-D-Elektrophoresetechniken eingesetzt.

Zu Beginn der Untersuchungen zeigte ein Vergleich mit Kontroll- und Patientenserum, daß beide Serumgruppen erdnußspezifische IgG-, IgA- und IgM-Antikörper enthielten. Dagegen wurden spezifische IgE-Antikörper nur mit dem Patientenserum nachgewiesen. Im IEF-Immunoprint waren die intensivsten IgE-Bindungen im pH-Bereich von pH 5,5 bis 7,5 festzustellen und im SDS-PAGE-Immunoblot sind Hauptallergene in den Molekulargewichtsbereichen von 17, 30, 48–66 und 116 kD identifiziert worden.

Es konnten 8 Hybridomzellinien (PN-a bis PN-h), die monoklonale Antikörper gegen IgE-reaktive Einzelkomponenten aus dem Erdnußextrakt produzieren, etabliert werden. In ELISA-Inhibitionstests ließen sich identische Bindungsstellen der monoklonalen Antikörper und Patientenseren am Antigen nachweisen. Weiterhin wurde die Eignung der monoklonalen Antikörper zur Detektion versteckter Allergene in verarbeiteten Nahrungsmitteln geprüft.

Summary

The purpose of this study is to improve the diagnosis of peanut allergy. In order to standardize test substances for in vivo and in vitro diagnostic, the type I allergy-associated single components of peanuts have been identified and characterized with the aid of patients' sera and monoclonal antibodies. For allergen detection IEF-immunoprint-, SDS-PAGE-immunoblot- and 2-D electrophoresis-techniques have been used.

A comparison of control sera and patients' sera showed that both contained peanut specific IgG-, IgA- and IgM-antibodies. In contrast, peanut-specific IgE-antibodies were only detectable with patients' sera. In IEF-immunoprint the most intensive IgE-bindings showed up in pl-range from pH 5.5 to 7.5. In SDS-PAGE-immunoblot major allergens could be identified at molecular weight ranges of 17, 30, 48 to 66 and 116 kD.

Raising monoclonal antibodies against IgE-reactive components from peanut extract resulted in eight antibody-producing hybridoma cell lines, named PN-a to PN-h. ELISA-inhibition tests revealed common epitopes of monoclonal antibodies and patients' antibodies. Moreover, the monoclonal antibodies were tested to see whether they can be used for detection of hidden peanut allergens.

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Abbreviations

lg:

Immunglobulin

IEF:

Isoelektrische Fokussierung

pl:

Isoelektrischer Punkt

SDS-PAGE:

Natriumdodecylsulfat-Polyacrylamidgel-Elektrophorese

kD:

Kilo Dalton

ELISA:

Enzyme-linked-immunosorbent-assay

NC:

Nitrocellulose

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Authors and Affiliations

  1. Forschungsinstitut Borstel, FRG

    L. Uhlemann, W. -M. Becker & M. Schlaak

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  1. L. Uhlemann
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  2. W. -M. Becker
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  3. M. Schlaak
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Uhlemann, L., Becker, W.M. & Schlaak, M. Nahrungsmittelallergie: Identifizierung und Charakterisierung von Erdnußallergenen mit Patientenseren und monoklonalen Antikörpern. Z Ernährungswiss 32, 139–151 (1993). https://doi.org/10.1007/BF01614757

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