The Journal of Immunology, Vol 146, Issue 3 893-898, Copyright © 1991 by American Association of Immunologists

ARTICLES

Mapping the epitopes of neutralizing anti-human IL-3 monoclonal antibodies. Implications for structure-activity relationship

NA Lokker, U Strittmatter, C Steiner, B Fagg, P Graff, HP Kocher and G Zenke
Preclinical Research, Sandoz Pharma Ltd., Basle, Switzerland.

The epitopes of neutralizing mAb were mapped in order to identify a receptor binding site on human IL-3 (huIL-3). To initiate this structure and activity analysis, four neutralizing mAb were selected on the basis of preventing rhuIL-3 stimulated proliferation of peripheral blood cells from a patient with chronic myelogenous leukemia (CML). In order to identify continuous epitopes, the neutralizing mAb were assayed in a solid-phase ELISA for their reactivity with either denatured rhuIL-3 or with the peptides generated by digestion of rhuIL- 3 by using two different proteinases. Two of the neutralizing mAb recognized single fragments from both digestions. Amino acid (aa) sequence determination showed that these peptides overlap, defining a region of 22 aa (aa 29 to 50 of the mature rhuIL-3 protein). In a competition ELISA, the two continuous epitopes were shown to be linked to one another and to the two discontinuous epitopes, suggesting that all four neutralizing mAb bind to a discrete region of the IL-3 molecule, which might be involved in binding to the IL-3R.

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