| Ingredients | Stock concentration | Final concentration |
|---|---|---|
| Tris-HCl (pH 8.0) | 1 M | 30 mM |
| EGTA | 200 mM | 4 mM |
| Sucrose | - | 30% (w/v) |
| Protease inhibitor cocktail (cOmplete EDTA-free) | 50x (prepared according to manufacturer’s recommendation) | 2x |
| Pepstatin A | 1 mg/ml (in 100% methanol, see Note 5) | 20 μg/ml |
| DTT | 1 M | 6 mM |
| PMSF | 200 mM (in 100% ethanol) | 10 mM |
| Benzamidine | 320 mM | 10 mM |
Note: All stock solutions are prepared in ddH2O water if not stated otherwise. Stock solutions of protease inhibitor cocktail, pepstatin A and DTT stored at -20 °C; PMSF and Benzamidine stocks are prepared fresh. All of these are added to the buffer prior to the cell lysis step. It is important to carefully monitor the pH of this buffer. Deviations from pH 8.0 are detrimental for phagosome motility.