Figure 3

GFP and C-peptide expression in HIP:GFP transfected rES cells Rhesus ES cells were grown in differentiation media through stage 3 (bFGF). These cells, growing predominately as a monolayer on a glass coverslip, were transfected with the HIP:GFP construct using ExGen 500. Cells were allowed to differentiate in stage 3 media (bFGF) for 3 weeks, followed by 2 weeks in stage 4 media containing 1 nM Exendin 4. The cells were fixed in 3.4% formaldehyde and permeabilized with acetone. Cells were co-stained with DAPI and c-peptide. GFP expression was identified using a specific GFP filter and C-peptide detected by Tx-Red secondary antibody. In this field, two cells show both GFP and C-peptide expression (large arrows) and one cell expressing only C-peptide (small arrow).