Generation of the B and NK potentials was also dependent on BMP-4 + VEGF in the serum-free EB-forming culture
| . | ES Cell Line . | None . | 20* V† . | 1.5B‡ . | 1.5B2V . | 1.5B20V . | 15B . | 15B2V . | 15B20V . |
|---|---|---|---|---|---|---|---|---|---|
| 1st | A3-1 | 0 | 0 | (1)1-153 | 8 | 4 | 0 | 2 | 1 |
| 2nd | A3-1 | 0 | 0 | 1 | 7 | 4 | 0 | 2 | 2 |
| 3rd | E14 | 0 | (1)1-153 | 1 | 7 | 9 | 1 | ND1-155 | ND |
| 4th | E14 | 0 | 0 | 2 | 4 | 5 | 3 | ND | ND |
| . | ES Cell Line . | None . | 20* V† . | 1.5B‡ . | 1.5B2V . | 1.5B20V . | 15B . | 15B2V . | 15B20V . |
|---|---|---|---|---|---|---|---|---|---|
| 1st | A3-1 | 0 | 0 | (1)1-153 | 8 | 4 | 0 | 2 | 1 |
| 2nd | A3-1 | 0 | 0 | 1 | 7 | 4 | 0 | 2 | 2 |
| 3rd | E14 | 0 | (1)1-153 | 1 | 7 | 9 | 1 | ND1-155 | ND |
| 4th | E14 | 0 | 0 | 2 | 4 | 5 | 3 | ND | ND |
Cells harvested from EBs developed in the serum-free medium with or without factors were plated and cultured at 2 × 104cells/well on a confluent layer of OP9 cells as described in “Materials and methods.” On day 14 of the coculture, pre-B cell foci and LAK cell foci were inspected under the microscope. Total lymphocyte foci numbers obtained from four independent experiments are shown. Then the nonadherent cells and loosely attached cells were harvested as described in “Materials and methods,” and the surface expressions B220, CD19, and Sca-1 were analyzed by FACScan as described.17
The value corresponds to concentration (ng/mL) of factor added during the EB formation.
V stands for VEGF; 2V and 20V mean 2 ng/mL and 20 ng/mL, respectively, of VEGF were added during the EB formation.
B stands for BMP-4; 1.5B and 15B mean 1.5 ng/mL and 15 ng/mL, respectively, of BMP-4 were added during the EB formation.
According to our previous results, ES cell-derived LAK cells are B220+ CD19−, and pre-B cells are B220+ CD19+.17 There were only a few cells in this well, and they were not B220+.
Not determined.