Properties of a surface antigen expressed on activated human thymus-derived lymphocytes

We have identified previously a quantitatively minor membrane protein (p28) with an apparent reduced m.w. of 28,000, which is biosynthetically labeled in activated human lymphocytes. Rabbit antisera with activity directed against p28 (alpha-ATC) were prepared and p28 was identified by immunoprecipitation in NP-40 extracts of activated, extrinsically labeled lymphocytes. p28 was not expressed in appreciable amounts by unstimulated T cells, stimulated or unstimulated B cells, null cells, or adherent cells. Protein p28 was only minimally represented on resting thymocytes but was easily detected on 4-hr activated thymocytes and the T lymphoblastoid cell lines HSB2 and MOLT-4. Absorption and immunoprecipitation studies with alpha-ATC indicated that p28 was not present on erythrocytes, platelets, neutrophils, six B cell lines, six null cell lines, and seven other T lymphoblastoid cell lines. Protein p28 from HSB2 cells was absorbed by lentil lectin, concanavalin A, and wheat germ agglutinin affinity columns and was eluted with the appropriate sugars. Gel filtration column chromatography of unreduced p28 in the presence of 0.5% NP-40 or 0.1% deoxycholate gave elution characteristics consistent with a m.w. of approximately 60,000 to 100,000. In preparative isoelectric focusing (IEF) studies the isoelectric point (pI (p28) = 5.2 to 6.1) was similar or identical to that described for the reduced and denatured protein in two-dimensional polyacrylamide gels (pI = 5.5 to 6.2). Protein p28 was eluted from DEAE-cellulose (Whatman DE-52) ion exchange columns at 0.05 to 0.15 M NaCl. Experiments with monoclonal antibodies or heteroantisera specific for other T cell and B cell antigens and various lymphoblastoid cell lines and normal peripheral blood cells indicated that p28 is distinct from the human Ia-like antigens, from T3, T4, T5, T8, and from several other reported human T cell antigens that appear to correspond to Thy-1, the sheep erythrocyte receptor, and a human thymus-leukemia antigen.