To support its ongoing monitoring program of therapeutic and diagnostic allergen preparations, LIB has initiated a multifaceted study of the factors that enhance or diminish the immune responses to allergens. These include: 1. Lipopolysaccharide (LPS) effect on allergen responses. LPS is a common contaminant in latex gloves and in the environment. LPS is also present in many allergen vaccines. In previous work, Dr. Slater has demonstrated that LPS increases murine IgG and IgE responses to recombinant Hev b 5, that this effect can be observed with ovalbumin as well, and that the effect is enhanced by lower airway delivery of the allergen and LPS. Current work is focused on two areas of interest. Ms. Valerio is examining the cytokine specificity of the LPS-allergen responses, and will identify the anatomic localization of the murine airway responses to LPS. We are also measuring the endotoxin content of commercial allergen vaccines, and will determine the effect of the endotoxin in these vaccines on murine responses to allergen injection. 2. Hev b 5 epitope study. Hev b 5 is an important allergen in Hevea latex products. The identification of relevant epitopes can be used as a tool for the immunomodulation of responses to latex allergens. Dr. Slater has identified putative B-cell and T-cell epitopes of Hev b 5 in BALB/c mice, and has developed mutants at some of these sites. Specific mutants, including proteins that has been mutated at 2 or 3 putative sites, have been expressed. Antibody binding studies have confirmed the importance of some of these sites. Antibody studies have been performed using monoclonal antibodies obtained from Drs. O'Hehir and Sutherland. These studies are being done by Ms. Patterson. 3. Cross-reactive allergens in foods and latex. Cross-reactivity among allergens can provide important insights into the relationships of structure to immunogenicity. Dr. Hsieh characterized and sequenced a major allergen in avocado, Prs a 1, which has chitinase activity an cross-reacts with the latex allergen Hev b 6. Dr. Poley has initiated studies to identify the effect of pre-sensitization with cross-reactive fruits on latex immunity. 4. Allergen vaccines are the most commonly utilized reagents for immunotherapy, and the standardization of allergens is an important step in ensuring the safety and efficacy of allergens for immunotherapy. Dr. Slater and Ms. Patterson have initiated work on the standardization of cockroach allergens in the US. In initial work, Ms. Patterson has demonstrated that currently available cockroach allergen vaccines are highly variable in their protein and allergen content. 5. Lyophilization of allergen vaccines. Dr. Slater, Ms. Valerio and Mr. Alston have shown that the lyophilization of allergen vaccines does not affect their ability to bind antibody in standard assays, and does not affect SDS-PAGE electrophoretic patterns significantly. Limited circular dichroism spectroscopy of cat allergen vaccines, performed and analyzed by Dr. Dixon, has indicated that lyophilization has no effect on secondary structure as well. 6. Construction of a combinatorial library for cockroach allergen antibodies. The study of cockroach allergens requires an analysis of the immunodominant proteins within a highly complex mixture. Dr. Finlay is using recombinant DNA technology to construct a library of human antibodies to cockroach. This technology will allow us to identify and study relevant allergens.
