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Proteomics Core |
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| Labeling Compatibility Information | |
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| A number of reagents commonly used for 2D gel electrophoresis have been tested to determine their impact on protein labeling using the DIGE approach. The following list details those reagents known to affect Cy dye labeling of proteins. The amount listed for each reagent is the maximum amount allowable. | |
| Reagent | Amersham Recommendations | Core Results |
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Dithiothreitol (DTT) |
2 mg/mL slight reduction in labeling | 2 mg/mL Significant reduction in labeling |
| Tris-(2-carboxyethyl) phosphene (TCEP) | 0.5-1mM - slight reduction in labeling | Not tested |
| b-mercaptoethanol | not recommended | Not tested |
| Triton X-100 | 1% causes 17% drop in labeling | Not tested |
| NP-40 | up to 1% | Not tested |
| SDS | up to 1% | Not tested |
| Tris | 10-40 mM (pH 8.0-9.0) | 30 mM (pH 8-9) OK |
| HEPES | not recommended | Not tested |
| Bicarbonate | 5 mM (pH 8.5) | 50 mM pH 8 |
| CHES | 5 mM (pH 9-9.5) | Not tested |
| PPA | 5 mM (pH 8.0) causes drop in labeling | Not tested |
| Pefabloc (AEBSF) | not recommended | Not tested |
| Pefabloc SC Plus | compatible at manufacturer's recommended conc. | OK |
| Complete Protease inhibitor | not recommended | OK |
| APMSF | compatible at manufacturer's recommended conc. | Not tested |
| EDTA | compatible at 0.5-1.0 mM | 5 mM OK |
| PMSF | compatible at manufacturer's recommended conc. | Decreases labeling |
| PMSF Plus | compatible at manufacturer's recommended conc. | OK |
| Pepstatin A | compatible at manufacturer's recommended conc. | OK |
| Phosphatase inhibitor cocktail 1 (Sigma) | compatible at manufacturer's recommended conc. | OK |
| Phosphatase inhibitor cocktail 2 (Sigma) | compatible at manufacturer's recommended conc. | OK |