In vitro differentiation of TER-119+/4A5+ cells cultured at limiting dilution5-150
| Time and Factor in Culture . | Total Cell Number . | Benzidine+ . | ACHE+ . |
|---|---|---|---|
| 0 h | 100 | 0 | 0 |
| 48 h, −GF | 23 ± 9 | 10 ± 3 | < 1 |
| 48 h, +EPO | 58 ± 16 | 45 ± 12 | 4 ± 2 |
| 48 h, +TPO | 55 ± 12 | 30 ± 10 | 13 ± 5 |
| Time and Factor in Culture . | Total Cell Number . | Benzidine+ . | ACHE+ . |
|---|---|---|---|
| 0 h | 100 | 0 | 0 |
| 48 h, −GF | 23 ± 9 | 10 ± 3 | < 1 |
| 48 h, +EPO | 58 ± 16 | 45 ± 12 | 4 ± 2 |
| 48 h, +TPO | 55 ± 12 | 30 ± 10 | 13 ± 5 |
TER-119+/4A5+ cells (isolated by cell sorting, 80%-83% pure) were plated at a theoretical concentration of 100 cells/well in 96 flat-bottomed plates containing IMDM supplemented with Nutridoma and incubated for 48 hours without growth factors (GFs) or in the presence of either erythropoietin (EPO; 5 U/mL) or thrombopoietin (TPO; 100 ng/mL) (4 wells per experimental point). After 48 hours of incubation, the plates were centrifuged and the medium carefully removed; cells were air-fixed at the bottom of the plate and stained with either benzidine or acetylcholinesterase as described in “Materials and methods.” The results are presented as the mean (± SD) of 3 separate experiments.