Abstract Eosinophils (EOSs) cultured in the presence of 50% peripheral blood mononuclear cell (PBMC)-derived culture supernatants remained 67% ± 7% (mean ± SEM; n = 5) viable for 7 days. In the absence of PBMC supernatant, only 15% ± 7% of cells remained viable for 7 days. PBMC supernatants from six atopic individuals, with eosinophilia, and six normal subjects, with no eosinophilia, were compared for EOS viability-enhancing activity with the same target EOSs. Optimal conditions for the production of viability-enhancing activity by mononuclear cells were established as a 24-hour culture period, with a concentration of 2 × 10 6 cells per milliliter. Comparison of monocyte-enriched and lymphocyte-enriched culture supernatants for the production of the EOS viability-enhancing activity indicated that both cell types released the factor. C-18 Sep-Pak separation of the PBMC culture supernatant yielded a major EOS viability-enhancing activity in the aqueous eluent, suggesting a hydrophilic molecule. This major activity was neutralized by a specific antibody to granulocyte/macrophage colony-stimulating factor but was unaffected by specific antibodies to interleukin-3 and interleukin-5. A second, minor viability-enhancing activity was observed in the 100% methanol fraction, indicating the presence of a more hydrophobic molecule. The supernatants from the PBMCs of the atopic individuals consistently enhanced EOS survival to a greater extent than supernatants from the PBMCs of the normal, nonatopic individuals.