CHARACTERIZATION OF ANTIGEN-SPECIFIC T CELL PRODUCTS

我们拟分离并鉴定抗原b特异性T辅助细胞（THF）

We propose to isolate and characterize antigen-B-specific T helper (THF) and T suppressor (TSF) factors from either murine spleen cell populations or appropriate T cell lines or hybridomas. The amount of THF and TSF produced will be quantitated by inhibition of monoclonal anti-AgB antibody binding to AgB bound to microtiter plates (ELISA). Methods used for factor purification will include affinity chromatography, gel-filtration, ion-exchange chromatography, isoelectrofocusing, and high pressure liquid chromatography (HPLC). The purified THF and TSF factors will be used to determine the relationship with immunoglobulin molecules that share antigen-binding properties but otherwise appear structurally unrelated, the nature of the interaction between the antigen-binding and I-region chains, and the role of the carbohydrate moiety in our factors. Another important goal is to develop an immunotherapy model with which to determine the potential of AgB-specific TSF for the treatment of timothy-sensitive patients. These studies will allow us to determine the optimal dose and frequency of TSF treatment to achieve maximal suppression of timothy IgE formation, and to determine the effect of multiple doses of TSF upon the IgE response of hyperimmunized animals. Anti-idiotypic antibodies (anti-THF and anti-IgEid) passed over Sepharose-fetal calf serum, -normal mouse serum, and -F(ab)2 adsorbents and finally eluted from Sepharose-TSF adsorbents will be used to determine the relationship between the idiotype expressed on T and B cells involved in IgG and IgE formation to the same antigenic determinant. Experiments to develop a tissue culture method to study the in vitro synthesis of murine IgE will permit us to examine the mechanism of IgE regulation by isolated T cell factors (THF, TSF1, and TSF2), and examine the site of action of these factors. The IgE recovered from future supernatants will be assayed by an ELISA method using a fluorescent substrate.