Percentage of chimerism after intrathymic injection of sorted embryonic bone marrow cells
| Injected cells per thymic lobe . | Number of animals . | % of chimerism in host thymus . |
|---|---|---|
| Unselected BM cells | ||
| 6 × 103 | 3 | 0 |
| 104 | 2 | 0 |
| 2 × 104 | 4 | 4 ± 2 |
| Cells stained with RR5 and c-kit* | ||
| 103c-kit+/RR5+ | 12 | 13.7 ± 1.8 |
| 102 | 8 | 2.4 ± 0.8 |
| 30 | 8 | 0.9 ± 0.2 |
| 10 | 4 | 1.7 ± 0.8 |
| 3 | 4 | 0.6 ± 0.3 |
| 103c-kit+/RR5− | 5 | 2.3 ± 1 |
| Cells stained with either c-kit or RR5† | ||
| 103Rhhi/c-kit+ | 3 | 6.6 ± 2.5 |
| 103Rhhi/c-kit− | 3 | 0.6 ± 0.4 |
| 103Rhhi/RR5+ | 3 | 15.0 ± 8.2 |
| 103Rhhi/RR5− | 3 | 2.7 ± 1.2 |
| Injected cells per thymic lobe . | Number of animals . | % of chimerism in host thymus . |
|---|---|---|
| Unselected BM cells | ||
| 6 × 103 | 3 | 0 |
| 104 | 2 | 0 |
| 2 × 104 | 4 | 4 ± 2 |
| Cells stained with RR5 and c-kit* | ||
| 103c-kit+/RR5+ | 12 | 13.7 ± 1.8 |
| 102 | 8 | 2.4 ± 0.8 |
| 30 | 8 | 0.9 ± 0.2 |
| 10 | 4 | 1.7 ± 0.8 |
| 3 | 4 | 0.6 ± 0.3 |
| 103c-kit+/RR5− | 5 | 2.3 ± 1 |
| Cells stained with either c-kit or RR5† | ||
| 103Rhhi/c-kit+ | 3 | 6.6 ± 2.5 |
| 103Rhhi/c-kit− | 3 | 0.6 ± 0.4 |
| 103Rhhi/RR5+ | 3 | 15.0 ± 8.2 |
| 103Rhhi/RR5− | 3 | 2.7 ± 1.2 |
BM = bone marrow.
Cells were sorted by fluorescence activated cell sorter (FACS), diluted according to FACS cell count, and 10 μL were injected per thymic lobe (8 lobes injected per chick). At concentrations below 100 cells per 10 μL, 1000 double negative cells were added as “carriers.” Control experiments with 4000 double negative cells resulted in a 0.14% chimerism, which is within the range of the accuracy of the method.
Cells were subsequently incubated with rhodamine 123 (Rh), FACS sorted, and injected intrathymically. Chimerism of the host thymus (in a pool of 4 lobes per chick) was determined 14 days later after staining for the ov marker and exclusion of dead cells by propidium iodide.