IgE mediated release of 3H-5HT from human mast cells (LAD cells)

Abstract

RATIONALE: The availability of human mast cell lines offers the possibility that antigen-specific IgEs in body fluids may be examined for their ability to activate human mast cells. The confounding factor is that β-hexosaminidase (β-hex) and histamine assays are difficult to employ because of the natural presence of these mediators in serum. We therefore determined whether the LAD human mast cell lines would take up serotonin (3H-5HT) and release this molecule following FcεRI cross linking. METHODS: LAD cells were incubated with 3H-5HT, followed by sensitization with biotinylated IgE (bIgE) and cross linking with streptavidin (SA). 3H-5HT release was compared with β-hex release. In some experiments, cells were incubated with dialyzed serum from ragweed sensitive individuals followed by activation with ragweed antigen E. In other experiments, cells were sensitized with bIgE in the presence of serum known to contain IgE by RAST and checked for release after SA cross linking. RESULTS: Uptake of 3H-5HT by mast cells reached a plateau by 48 hours. Percent IgE mediated 3H-5HT release paralleled % β-hex release. Highest % release for LAD cells was 60%. Release from LAD cells using 1:25-1:50 dilutions of dialyzed ragweed serum followed by the addition of ragweed antigen E reached over 21%. LAD cells could be sensitized in the presence of human sera. CONCLUSIONS: Human mast cells may be used in a 3H-5HT release assay to measure activation with bIgE/SA or human serum from ragweed sensitive individuals. The presence of serum containing IgE does not inhibit bIgE-mediated release of 3H-5HT.