A method for differentiating human pluripotent stem cells into cells expressing markers characteristic of the pancreatic endocrine lineage, comprising the steps of: a. Culturing the human pluripotent stem cells on a tissue culture substrate coated with an extracellular matrix, wherein the extracellular matrix is selected from the group consisting of a solubilized basement membrane preparation extracted from mouse sarcoma cells, growth factor-reduced solubilized basement membrane preparation extracted from mouse sarcoma cells, laminin, fibronectin, and human serum; b. Differentiating the human pluripotent stem cells into cells expressing markers characteristic of the definitive endoderm lineage, by treating the human pluripotent stem cells with 1 pg/ml to 100 μg/ml of activin A and 1 ng/ml to 1000 ng/ml of Wnt-3a in a first culture medium for 1 to 3 days, followed by culturing the human pluripotent stem cells with 1 pg/ml of activin A and optionally 1 ng/ml to 1000 ng/ml of Wnt-3a in a second culture medium for 1 day to 4 days; c. Differentiating the cells expressing markers characteristic of the definitive endoderm lineage into cells expressing markers characteristic of the pancreatic endoderm lineage, by treating the cells expressing markers characteristic of the definitive endoderm lineage with 1 nM to 1 mM of retinoic acid and 50 pg/ml to 50 μg/ml of at least one fibroblast growth factor selected from the group consisting of FGF-2, FGF-4 and FGF-10 for 1 to 3 days; and d. Differentiating the cells expressing markers characteristic of the pancreatic endoderm lineage into cells expressing markers characteristic of the pancreatic endocrine lineage, by treating the cells expressing markers characteristic of the pancreatic endoderm lineage with 0.1 μM to 100 μM of L-685,458 for 3 to 5 days; wherein step b. optionally further comprises treating the cells with 1 nM to 1000 nM of a GSK-3B inhibitor, and wherein the GSK-3B inhibitor is a GSK-3B inhibitor XI or GSK-3B inhibitor IX. a. Culturing the human pluripotent stem cells on a tissue culture substrate coated with an extracellular matrix, wherein the extracellular matrix is selected from the group consisting of a solubilized basement membrane preparation extracted from mouse sarcoma cells, growth factor-reduced solubilized basement membrane preparation extracted from mouse sarcoma cells, laminin, fibronectin, and human serum; b. Differentiating the human pluripotent stem cells into cells expressing markers characteristic of the definitive endoderm lineage, by treating the human pluripotent stem cells with 1 pg/ml to 100 μg/ml of activin A and 1 ng/ml to 1000 ng/ml of Wnt-3a in a first culture medium for 1 to 3 days, followed by culturing the human pluripotent stem cells with 1 pg/ml of activin A and optionally 1 ng/ml to 1000 ng/ml of Wnt-3a in a second culture medium for 1 day to 4 days; c. Differentiating the cells expressing markers characteristic of the definitive endoderm lineage into cells expressing markers characteristic of the pancreatic endoderm lineage, by treating the cells expressing markers characteristic of the definitive endoderm lineage with 1 nM to 1 mM of retinoic acid and 50 pg/ml to 50 μg/ml of at least one fibroblast growth factor selected from the group consisting of FGF-2, FGF-4 and FGF-10 for 1 to 3 days; and d. Differentiating the cells expressing markers characteristic of the pancreatic endoderm lineage into cells expressing markers characteristic of the pancreatic endocrine lineage, by treating the cells expressing markers characteristic of the pancreatic endoderm lineage with 0.1 μM to 100 μM of L-685,458 for 3 to 5 days; wherein step b. optionally further comprises treating the cells with 1 nM to 1000 nM of a GSK-3B inhibitor, and wherein the GSK-3B inhibitor is a GSK-3B inhibitor XI or GSK-3B inhibitor IX.