BD StemflowTM Human Neural Lineage Analysis Kit

Description

Neural Induction of H9 hESC: H9 Human embryonic stem cells hESC Wicell Madison, WI were cultured on mouse embryonic fibroblasts in hESC medium DMEM:F12, 20 % Knockout Serum Replacement KOSR , non-essential amino acids, 20 mM glutamine Invitrogen , 1X penicillin/streptomycin Lonza , bFGF Cat. No. 354060 . Cell cultures were treated with Dispase Cat. No. 354235 and cell colonies were scraped and placed in hESC medium without FGF in low adhesion plates to form embroid bodies EB . After 4 days the EBs were plated on BD Matrigel-coated plates Cat. No. 354277 and placed in neural induction medium DMEM:F12 Invitrogen , 1X ITS supplement Sigma , 250 ng/ml recombinant Noggin R&D Systems and penicillin/streptomycin Lonza and cultured for an additional 15 days. Differentiation of H9-derived NSC: H9 hESC-derived NSC were differentiated in neural differentiation medium DMEM:F12, 0.5X B27, 0.5X N2 Invitrogen , 1X penicillin/streptomycin Lonza , 20 ng/ml BDNF, 20ng/ml GDNF both from Peprotech and 0.5 mM dibutyryl cyclic AMP Sigma for 28 days. Differentiation of H9-derived NSC: H9 hESC-derived NSC were differentiated in neural differentiation medium DMEM:F12, 0.5X B27, 0.5X N2 Invitrogen , 1X penicillin/streptomycin Lonza , 20 ng/ml BDNF, 20ng/ml GDNF both from Peprotech and 0.5 mM dibutyryl cyclic AMP Sigma for 11 days. Culture medium was replaced with AGM Astrocyte Growth Medium Lonza for an additional 16 days. Neural Induction of H9 hESC: H9 Human embryonic stem cells hESC Wicell Madison, WI were cultured on mouse embryonic fibroblasts in hESC medium DMEM:F12, 20 % Knockout Serum Replacement KOSR , non-essential amino acids, 20 mM glutamine Invitrogen , 1X penicillin/streptomycin Lonza , bFGF Cat. No. 354060 . Cell cultures were treated with Dispase Cat. No. 354235 and cell colonies were scraped and placed in hESC medium without FGF in low adhesion plates to form embroid bodies EB . After 4 days the EBs were plated on BD Matrigel-coated plates Cat. No. 354277 and placed in neural induction medium DMEM:F12 Invitrogen , 1X ITS supplement Sigma , 250 ng/ml recombinant Noggin R&D Systems and penicillin/streptomycin Lonza and cultured for an additional 15 days. Differentiation of H9-derived NSC: H9 hESC-derived NSC were differentiated in neural differentiation medium DMEM:F12, 0.5X B27, 0.5X N2 Invitrogen , 1X penicillin/streptomycin Lonza , 20 ng/ml BDNF, 20ng/ml GDNF both from Peprotech and 0.5 mM dibutyryl cyclic AMP Sigma for 28 days. Show More Differentiation of H9-derived NSC: H9 hESC-derived NSC were differentiated in neural differentiation medium DMEM:F12, 0.5X B27, 0.5X N2 Invitrogen , 1X penicillin/streptomycin Lonza , 20 ng/ml BDNF, 20ng/ml GDNF both from Peprotech and 0.5 mM dibutyryl cyclic AMP Sigma for 11 days. Culture medium was replaced with AGM Astrocyte Growth Medium Lonza for an additional 16 days. Show More Neural Induction of H9 hESC: H9 Human embryonic stem cells hESC Wicell Madison, WI were cultured on mouse embryonic fibroblasts in hESC medium DMEM:F12, 20 % Knockout Serum Replacement KOSR , non-essential amino acids, 20 mM glutamine Invitrogen , 1X penicillin/streptomycin Lonza , bFGF Cat. No. 354060 . Cell cultures were treated with Dispase Cat. No. 354235 and cell colonies were scraped and placed in hESC medium without FGF in low adhesion plates to form embroid bodies EB . After 4 days the EBs were plated on BD Matrigel-coated plates Cat. No. 354277 and placed in neural induction medium DMEM:F12 Invitrogen , 1X ITS supplement Sigma , 250 ng/ml recombinant Noggin R&D Systems and penicillin/streptomycin Lonza and cultured for an additional 15 days. Differentiation of H9-derived NSC: H9 hESC-derived NSC were differentiated in neural differentiation medium DMEM:F12, 0.5X B27, 0.5X N2 Invitrogen , 1X penicillin/streptomycin Lonza , 20 ng/ml BDNF, 20ng/ml GDNF both from Peprotech and 0.5 mM dibutyryl cyclic AMP Sigma for 28 days. Differentiation of H9-derived NSC: H9 hESC-derived NSC were differentiated in neural differentiation medium DMEM:F12, 0.5X B27, 0.5X N2 Invitrogen , 1X penicillin/streptomycin Lonza , 20 ng/ml BDNF, 20ng/ml GDNF both from Peprotech and 0.5 mM dibutyryl cyclic AMP Sigma for 11 days. Culture medium was replaced with AGM Astrocyte Growth Medium Lonza for an additional 16 days.