Quick Start Protocol for NEBExpress® Ni-NTA Magnetic Beads
- Gently vortex and thoroughly suspend magnetic beads.
- Aliquot 50 μl of bead suspension to a sterile microcentrifuge tube, apply magnet, to pull beads to the side of the tube and remove supernatant.
- Equilibrate beads by mixing with 200 μl of binding buffer (20 mM sodium phosphate, 300 mM NaCl, 10 mM Imidazole, pH 7.4), apply magnet to pull the beads aside and remove the supernatant.
- Add 0.2-1 ml cell extract and incubate with mixing at 4°C for 30 minutes. Apply magnet and remove supernatant.
- Wash beads 3X with 500 μl of wash buffer (20 mM sodium phosphate, 300 mM NaCl, 20 mM Imidazole, pH 7.4) as per Step 3.
- Elute fusion protein by incubating with 100 μl elution buffer (20 mM sodium phosphate, 300 mM NaCl, 500 mM Imidazole, pH 7.4) for ≥ 2 minutes at 4°C with mixing. Apply magnet, remove and keep supernatant. Elution can be repeated, and eluates combined.