A mitogenic heparin-binding (reactive) lectin-like protein (HBP) was purified from the extract of a cloned rat thymic myoid cell R615B2 by a one-step procedure of affinity chromatography on a heparin--Sepharose CL-6B column. Four distinct peptide bands with molecular weights of 10,000, 13,000, 13,700, and 14,600 were detected on SDS-polyacrylamide gel electrophoresis. This protein is mitogenic at concentrations of as low as 1.1-70.0 ng/ml for peanut lectin-nonagglutinated thymocytes and splenocytes from euthymic mice and rats but not for splenocytes from nude mice. These results indicate that thymic myoid cell-derived HBP is an important signal for one particular step in T-cell differentiation.