Feeder cells from different sources and conditioned media for recloning of human--mouse and mouse--mouse hybridomas

The use of different feeder cell layers (peritoneal macrophages from mice and rats, spleen cells and thymocytes from mice) for recloning of human--mouse and mouse--mouse hybridomas has been described. Optimal numbers of feeder cells from different sources required for high cloning efficiencies were determined. It was possible to use cryopreserved rat and mouse macrophages as feeders for cell cloning. However, the resulting cloning efficiency was much lower in comparison to the fresh material. Culture supernatants from human endothelial cells (added in a final concentration of 40% v/v) and from chicken embryo fibroblasts (25%) could replace the feeder cell layer in recloning experiments with both human--mouse and mouse--mouse hybridomas. Therefore, conditioned media (prepared in large quantities) may be used for generating standardized conditions for high-efficient cloning and recloning of hybridoma cell lines.