TrkA tyrosine phosphorylation in response to MAb 5C3
| Cells | C10 Cells | 4-3.6 Cells | |||
|---|---|---|---|---|---|
| PY total | PY490 | PY total | PY490 | ||
| 1 | No ligand | 11 | 1 | 4 | 1 |
| 2 | NGF 1 pm | 12 | 1 | 7 | 5 |
| 3 | NGF 10 pm | 12 | 1 | 44 | 33 |
| 4 | NGF 100 pm | 36 | 45 | 93 | 61 |
| 5 | NGF 1 nm | 100 | 100 | 100 | 100 |
| 6 | 5C3 0.05 nm | 10 | 1 | 5 | 1 |
| 7 | 5C3 0.5 nm | 40 | 40 | 32 | 21 |
| 8 | 5C3 5 nm | 91 | 71 | 45 | 43 |
| 9 | 5C3 50 nm | 35 | 49 | 39 | 21 |
Cells were untreated (lane 1) or treated with the indicated concentrations of NGF (lanes 2–5) or mAb 5C3 (lanes 6–9), for 15 min at 37°C. Ligand concentrations were selected on the basis of survival assays (e.g., Table 3). Equal amounts of protein from whole cell lysates were resolved by SDS-PAGE and analyzed by Western blotting with antiphosphotyrosine (anti-PY) or with α-PY490 blot (DF-49 sera) recognizing specifically the Shc binding site of TrkA. Band intensities were analyzed by densitometry and standardized using the relative optical density of 1 nm NGF treatment as 100%. Data from a representative Western blot are shown.