PRODUCTION, SPECIFICITY, AND FUNCTIONALITY OF MONOCLONAL-ANTIBODIES TO SPECIFIC PEPTIDE-MAJOR HISTOCOMPATIBILITY COMPLEX CLASS-II COMPLEXESFORMED BY PROCESSING OF EXOGENOUS PROTEIN

Several unanswered questions in T cell immunobiology relating to intra cellular processing or in vivo antigen presentation could be approache d if convenient, specific, and sensitive reagents were available for d etecting the peptide-major histocompatibility complex (MHC) class I or class II ligands recognized by alpha beta T cell receptors. For this reason, we have developed a method using homogeneously loaded peptide- MHC class II complexes to generate and select specific mAb reactive wi th these structures using hen egg lysozyme (HEL) and I-A(k) as a model system. mAbs specific for either HEL-(46-61)-A(k) or HEL-(116-129)-A( k) have been isolated. They cross-react with a small subset of I-A(k) molecules loaded with serf peptides but can nonetheless be used for fl ow cytometry, immunoprecipitation, Western blotting, and intracellular immunofluorescence to detect specific HEL peptide-MHC class II comple xes formed by either peptide exposure or natural processing of native HEL. An example of the utility of these reagents is provided herein by using one of the anti-HEL-(46-61)-A(k) specific mAbs to visualize int racellular compartments where I-A(k) is loaded with HEL-derived peptid es early after antigen administration. Other uses, especially for in v ivo tracking of specific ligand bearing antigen presenting cells, are discussed.