Localization of Thymosin α 1 Production to Thymus Medullary Epithelial Cells by Use of Monoclonal Antibodies

Abstract

The thymus is an endocrine organ which modulates T-cell immunity through the production of protein like peptides such as the thymosins. Thymosin α₁ was the first biologically active peptide isolated and sequenced from the partially purified thymic preparation, thymosin fraction 5, and has been extensively studied. Using synthetic Thymosin α₁, a heterologous rabbit antiserum has been raised and a radioimmunoassay has been developed. Although thymosin α₁ antibodies have been used in several histological studies, their use is limited by potential nonspecific cross-reactivities, unpredictable heterogenicity, variable affinities, and a limited unstandardized supply. In the studies, reported here, eight anti-thymosin α₁ monoclonal antibodies (MAbs) were produced by somatic cell fusion between spleen cells from immunized BALB/c mice and P3x64 Ag8.653 myeloma cells. The MAbs were screened for anti-thymosin α₁ specificity in a solid phase ELISA and a liquid phase RIA. Only those clones which secreted specific antibody as detected by both procedures were characterized for their heavy chain class and epitope specificity.

The anti-thymosin α₁ monoclonal antibodies were then used for indirect immune fluorescence studies of perfused rat thymus. Thymosin α₁ containing cells were found primarily in the thymic medulla, confirming previous studies using the heterologous antisera. These studies demonstrated the specificity of the anti-thymosin α₁ monoclonal antibodies for immunochemical studies of intra-and extra-thymic localization of thymosin α₁. They also provide an important reagent for biological studies of the role of thymosin α₁, in vitro and in vivo.