Table 4.
Plates with their contents used for the automated phosphopeptide enrichment on KingFisher Flex or Apex models
| KingFisher 96 deep well plate order | |||
|---|---|---|---|
| Plate number and name | μL/ well | Buffer | Time per step (min) |
| 1: Tip plate | |||
| 2: MagReSyn Ti-IMAC HP or Zr-IMAC HP beads (1:2 ratio bead to peptide) | Varies based on peptide amount Table 5 | 20% EtOH | |
| 3: Microsphere equilibration | 500 | Phosphopeptide Bind buffer | 5 |
| 4: Peptides (or TMT-labeled peptides) | 200 | Phosphopeptide Bind buffer | 20 |
| 5: Wash | 500 | Phosphopeptide Bind buffer | 2 |
| 6: Wash | 500 | Phosphopeptide Wash Buffer 1 | 2 |
| 7: Wash | 500 | Phosphopeptide Wash Buffer 2 | 2 |
| 8: Phosphopeptide elution | 200 | Phosphopeptide Elution Buffer | 10 |