Anti-mucin antibodies as tools in airway mucus characterization

The extensive release of mucus is a hallmark of many inflammatory airway disorders. In which respect and to what extent such inflammation-borne mucus deviates from the mucus that is released from the airway epithelium of nondiseased individuals has not been investigated in detail. In order to characterize the molecular properties of the main mucus constituent mucin in health and disease, we set out to develop monoclonal antibodies against the major human airway mucin MUC5AC. As the glycosylation pattern of mucosal surface constituents was shown to be highly diverse, even among individuals of the same species, we aimed to direct the antibodies against the protein backbone of humMUC5AC which ought to be better conserved than the carbohydrate antennae it carries. After an extensive analysis of the humMUC5AC protein sequence 4 peptide motifs were selected that neither should be covered by carbohydrates in the native protein nor display sequence homologies to other mucins. The peptides were coupled to carrier protein and the conjugates were used to immunize Balb/c mice. Strong polyclonal immune responses against the peptides were detected and for each of the selected sequences a battery of peptide-reactive monoclonal antibodies (moAbs) could be obtained. However, when testing the reactivity of the moAbs against native human nasal mucins obtained from different individuals it turned out that the antibodies did not exhibit a uniform recognition but rather showed a proband-dependent signal. Whether this is due to polymorphisms in the MUC5AC gene, i.e. mutations in the protein backbone, or to differences in MUC5AC expression rates or whether posttranslational or postsecretory modifications account for this result requires further investigation.