The presence of non-isotype-specific antibodies in polyclonal anti-IgE reagents: demonstration of their binding to specifically selected Epstein-Barr virus-transformed cell lines.
Polyclonal anti-human IgE reagents were earlier shown to contain variable amounts of nonisotype-specific antibodies depending on the strategy used for their preparation. The presence of these antibodies in two commercial anti-IgE reagents was demonstrated in this work by (a) their binding to human Ig-surface-positive lymphoblastoid cells specifically selected by one of the polyclonal anti-human IgE reagents and (b) their binding to the non-IgE immunoglobulins secreted by those lymphoblastoid cells. Peripheral blood B lymphocytes from two normal and two atopic patients were immortalized with Epstein-Barr virus (EBV) and then selected for cells that rosette with anti-IgE-coated erythrocytes. Selection was repeated four times and cells were then cloned. The cloned cells formed rosettes and their supernatants agglutinated erythrocytes coated with rabbit anti-IgE. The immunoglobulins of these clones were positive in an ELISA for IgE, using two different polyclonal anti-human IgE reagents. They were shown, however, to be 19 S IgMs. This discrepancy was due apparently to substantial contamination of anti-non-IgE-isotype-specific antibodies in the polyclonal anti-IgE reagents used both in the selection of cells and in the ELISA. The human monoclonal B-cell lines which were applied here as targets amplified the non-IgE-isotype specific antibody contamination present in the polyclonal anti-human IgE reagents. Because of the normally very low frequency of IgE-positive cells, the use of polyclonal anti-IgE reagents to detect these cells has to be carefully evaluated.
D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000276:immunology*; D000918:Antibody Specificity; D001666:Binding Sites, Antibody; D002461:Cell Line, Transformed; D002472:Cell Transformation, Viral; D004797:Enzyme-Linked Immunosorbent Assay; D006041:Goats; D004854:Herpesvirus 4, Human; D006801:Humans; D007073:Immunoglobulin E / Q000032:analysis / Q000276:immunology*; D007132:Immunoglobulin Isotypes / Q000032:analysis; D007075:Immunoglobulin M; D007202:Indicators and Reagents; D008213:Lymphocyte Activation; D011817:Rabbits; D012397:Rosette Formation; D013194:Staining and Labeling
D000888:Antibodies, Anti-Idiotypic; D007132:Immunoglobulin Isotypes; D007075:Immunoglobulin M; D007202:Indicators and Reagents; C026408:anti-IgE antibodies; D007073:Immunoglobulin E