Stimulating CD34+ hematopoietic cells with growth factors prior to transplantation may decrease the duration of post-transplant aplasia. The optimal time in which to deliver this stimulus remains unclear. We therefore sought to determine if progenitor cell cloning efficiency, as reflected by colony forming units-granulocyte-macrophage (CFU-GM) colony formation, differed when growth factor stimulation was carried out prior to freezing or after thawing. To address this issue, CD34+ marrow cells were suspended in Iscove's medium with 20% bovine calf serum. They were then stimulated with kit ligand (KL), interleukin-3 (IL-3), and interleukin-1 beta (IL-1 beta) for 36 h either prior to cryopreservation or immediately after thawing. We observed that cells stimulated prior to freezing formed more CFU-GM colonies than cells stimulated after thawing. We also found that CD34+ cells stimulated with growth factors either before or after freezing gave rise to more CFU-GM colonies than thawed cells plated without prestimulation. Thus, the cloning efficiency of stored marrow, as reflected by CFU-GM colony formation, may be effectively enhanced by pretreating cells with hematopoietic growth factors. Our data further suggest that the optimal time for this treatment is before cryopreservation as opposed to immediately after thawing.(ABSTRACT TRUNCATED AT 250 WORDS)