A limited collagenase digestion was used to prepare human fetal pancreatic (HFP) proislets that maintained in vitro and in vivo functional viability. Digestion times, collagenase concentrations, and culture conditions were optimized by assessing insulin release in response to low glucose (3.3 mM), high glucose (16.7 mM), and high glucose plus theophylline (10 mM). A stimulation index (ng insulin release in low glucose/ng insulin in high glucose plus theophylline) and insulin production (ng insulin/mg tissue) were used to determine functional viability. A media change 24 hr after digestion resulted in a marked increase in mean stimulation index (6.18± 1.2 vs. 3.12±0.9). IGF-1 was found to enrich β-cell viability as demonstrated by insulin-specific immuno- peroxidase staining and insulin release in response to HGT (36.3±7.7 ng/mg tissue vs. 18.1±5.2 ng/mg tissue). Immunohistologic staining of proislets suggested selective enrichment of β cells. Diabetic BALB/c/nu/nu transplanted with proislets cultured in the presence of IGF-1 (100 ng/ml) returned to normoglycemia in 6.7±3.1 weeks (range 4-11) versus 12.0±4.0 (range 10-15) in controls. Oral glucose tolerance test demonstrated in vivo serum glucose control equivalent to nondiabetic control mice. These findings suggest that HFP proislet preparation and culture with endocrine growth factors, specifically IGF-1, may provide a source of tissue with enriched β -cell activity that may have decreased immunogenic load and is more suitable for clinical transplantation. © 1991 by Williams & Wilkins.