Ni-NTA Superflow 96 BioRobot Kit (4)

Designed by QIAGEN for automated, medium-scale purification of His-tagged proteins, the Ni-NTA Superflow 96 BioRobot Kit provides automated purification of up to 300 µg recombinant protein from 96 samples in parallel using a standard protocol with a 5 ml culture volume per sample. Cleared bacterial cell lysates flow onto a filter plate preloaded with Ni-NTA Superflow resin by the BioRobot workstation. This 96-well metal-chelate affinity-chromatography module strongly and selectively binds His-tagged proteins, and ready-to-run protocols are provided for purification under native or denaturing conditions.

For larger culture volumes, high-yield protocols use 200 µl of Ni-NTA Superflow resin per well together with an optimized lysis buffer, enabling processing of up to 25 ml cultures under native conditions or 15 ml under denaturing conditions. Cell cultures are transferred to 24-well blocks, and up to 2 mg amounts of His-tagged protein per well can be purified. The kit is designed for use with BioRobot 3000, 8000, or 9600 workstations.

The system relies on patented Ni-NTA (nickel-nitrilotriacetic acid) resin, in which NTA provides four chelation sites for nickel ions — more than the three sites offered by other metal-chelating systems — preventing nickel-ion leaching and giving a higher binding capacity and purity. The single-step purification works under native or denaturing conditions from any expression system, including baculovirus, mammalian cells, yeast, and bacteria. A broad range of reagents is tolerated, including 6 M Gu·HCl, 8 M urea, 2% Triton X-100, 2% Tween 20, 1% CHAPS, 20 mM β-ME, 10 mM DTT, 20 mM TCEP, 50% glycerol, 20% ethanol, up to 30% ethanol, 5 mM CaCl₂ or 100 mM NaOH, 20 mM imidazole, 4 M MgCl₂, and 2 M NaCl. Purified proteins are eluted under mild conditions using 100–250 mM imidazole as competitor or by a reduction in pH. Key specifications include a yield range of 15 µg – 4 mg, bead size of 60–160 µm, binding capacity of 5–20 mg/ml, and 1–96 samples per run with automated processing.