6533b7d4fe1ef96bd12620da

RESEARCH PRODUCT

An approach to predictive testing of contact sensitizers in vitro by monitoring their influence on endocytotic mechanisms.

subject

description

Endocytotic activation of epidermal Langerhans cells (LC) by immunogenic haptens is an early event during development of allergic contact dermatitis. In this work a fast and objective flow-cytometric assay for predictive in vitro testing of contact sensitizers by monitoring their influence on endocytotic mechanisms in murine LC was developed. Epidermal cell suspensions were labelled with a monoclonal antibody directed to MHC class II molecules and pH-sensitive fluorochrome-coupled second-step reagents. For untreated LC a significant quenching of fluorescence intensity by internalization of the MHC-antibody complexes into acidic compartments was noticed. Similar results were obtained in the presence of irritants, the lectin concanavalin A and the phorbol ester phorbol 12-myristate 13-acetate. In contrast stimulation with several well-defined sensitizing compounds resulted in partial conservation of the fluorescence intensity due to the internalization of the labelled complexes into less acidic compartments. Monitoring this modulation of endocytosis is an effective in vitro method to test for properties typical for moderate and strong contact sensitizers. It will help to assess the risk of unknown chemicals to act as haptens and should be useful for restriction of animal experimentation in this field.