Evaluation of Murine Thymocyte Stimulation Using a Defined Culture Medium

Abstract

The ability of RPMI 1640 enriched with FCS or AATSZ (L-alanine, BSA, human transferrin, zinc chloride, and sodium selenite) to support mitogen-induced activation (G 0-G 1 shift) and proliferation (G 1-S shift) of thymocytes has been investigated. The two culture media were found to be equally supportive. In terms of viability, differences were detected in the number of recovered viable cells, but this could be related to alterations in adherent properties, rather than viability of the cells. For the examination of a PHA-induced proliferation, IL-1-containing suppernatants, deriving from normal or induced peritoneal macrophages, were prepared. The supportive capacity of these preparations showed no significant difference between AATSZ and FCS. Despite the excellent supportive capacity for the mitogen-stimulated thymocyte cultures, the AATSZ medium was not able to support all established cell lines tested. A T cell (MOLT 4F) and a macrophage cell line (SK 1) grew equally well in AATSZ- and FCS-enriched medium, but a B cell (U 266) and a null-cell line (Reh) did not proliferate at all. When cells from the latter two lines were cultured in AATSZ medium, they did not complete the RNA-synthesis required for DNA-synthesis, as judged by cytofluorography. From the experiments presented it is concluded that the AATSZ medium offers several advantages, such as easy standardization of culture conditions, and no essential disadvantages for studying mitogen-stimulated thymocytes in vitro. On the other hand, some lymphoid cell lines do require culture conditions that the AATSZ medium cannot provide.