Table 1.
Volumes of Buffers and Beads Used for Proximity Biotinylation and Protein Purification
| Pre-Incubation with BP | Biotinylation (PBS++ + H2O2) | STOP/Wash (×3) | Cell Lysis | Bead Slurry | Elution | |
|---|---|---|---|---|---|---|
| 100 mm dish | 7 mL | 12 mL | 12 mL | 1 mL | 50 μL | ~50 μL |
| 6-well platea | 2 mL | 4 mL | 4 mL | 500 μL | 10 μL | ~20 μL |
| 75 mm filter insert (Dish; Insert) | 7 mL; 7 mL | 10 mL; 10 mL | 8 mL; 8 mL | 1.2 mL | 50 μL | ~50 μL |
| 24 mm filter insert (Well; Insert) | 3 mL; 2 mL | 4 mL; 3 mL | 4 mL; 3 mL | 600 μL | 10 μL | ~20 μL |
a
For quenching of the reaction in multi-well plates, add 2× STOP solution to all wells to quench immediately, followed by two additional washes with 1× STOP solution.