PureCube Ni-NTA MagBeads

Order number: 31201

€87.00*

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Description

PureCube Ni-NTA magnetic beads / MagBeads were developed by Cube Biotech for His-tag protein purification. Our year-long experience in manufacturing agarose resin lead to the high yield of 80 mg protein per ml resin, which is leading in the market compared to other Ni-NTA suppliers.

Ni-NTA products by Cube Biotech are also available as agarose resin beads or as XL magnetic beads. For your convenience Cube Biotech also offers its own MagBead separator for use with our magnetic beads.
Feature
Usage Specific binding and purification of 6x His-tagged proteins
Specificity Affinity to His-tagged proteins
Binding capacity >80 mg/mL
Bead Ligand Ni-NTA
Bead size 30 μm
Filling quantity Delivered as a 25 % suspension
pH stability 2-14
Chelator stability Stable in buffer containing 10 mM DTT and 1 mM EDTA
Other stabilities 100% methanol, 100% ethanol, 8 M urea, 6 M guanidinium hydrochloride, 30% (v/v) acetonitrile

Lab Results

High yield and purity

Our unique production process yields a Ni-NTA Agarose that exhibits a protein binding capacity >20% higher than that of two leading competitor products. Figure 1 shows the SDS-PAGE of GFP expressed in E. coli and purified in gravity columns with PureCube Ni-NTA Agarose and the Ni-NTA resin from Competitor G and Competitor Q. The protein yield in 4 elutions (E1-E4, Cube) was 80 mg/mL, compared to 65 and 48 mg/mL obtained with the alternative resins (E1-E4, Competitor G, Competitor Q). Similar results (10-18% higher binding capacity; data not shown here) were obtained by comparing the purification of JNK1 (Kinase, 48 kDa) on PureCube Ni-NTA and the Ni-NTA of leading providers.
PureCube Ni-NTA compared to other competitors
Figure 1: Over 20% more yield was obtained with PureCube Ni-NTA Agarose. SDS-PAGE of GFP expressed in E. coli and purified in gravity columns with PureCube Ni-NTA Agarose and Ni-NTA resin from Competitor Q. 80 mg/mL protein yield was obtained with PureCube Ni-NTA Agarose (E1–E4, Cube) compared to 65 and 48 mg/mL, respectively, with the widely used alternative resins G and Q (E1–E4, Competitor G / Competitor Q).
Superior DTT and EDTA stability

PureCube Ni-NTA Agarose is very robust in the presence of DTT and EDTA. In a stability test, PureCube Ni-NTA Agarose was exposed to increasing concentrations of DTT or EDTA for 1 h. Thereafter, the resins were used to purify E. coli-expressed GFP-His in gravity columns. The binding capacity of the resin decreased in the presence of both DTT and EDTA but the decay rate was shallow. In presence of DTT, PureCube Ni-NTA Agarose lost on average 8% binding capacity with each increase in DTT concentration, resulting in an overall decay of 22% at 10 mM. Even at 1.5 mM EDTA, the resin still exhibits 54% of its maximum binding capacity (Fig. 2).
EDTA tolerance of PureCube Ni-NTA
Figure 2: NTA is robust in the presence of reducing and chelating agents. GFP-His was purified on gravity columns containing PureCube Ni-NTA Agarose after exposing the resin for 1 h to 3 concentrations of DTT or EDTA. NTA exhibits a shallow decay rate in binding capacity.
Robust against oxidation and regenerable

PureCube Ni-NTA Agarose retains its color and function after exposure to as much as 10 mM DTT. Figure 3 shows a photo series of the resin after a 1 h exposure to 5 mM DTT. Unlike other resins, PureCube Ni-NTA Agarose did not turn brown (A). The resin was still able to bind GFP (B), with a measured binding capacity of 65 mg/mL (see Fig. 2). The resin could then be regenerated by stripping the NTA, turning the resin white (C), and reloading it with nickel ions (D). The protocol for regenerating PureCube Ni-NTA Agarose can be downloaded.
EDTA tolerance of PureCube Ni-NTA
Figure 3: PureCube Ni-NTA Agarose is robust against oxidation and regenerable. PureCube Ni-NTA Agarose was exposed to 5mM DTT for 1 h (A). After demonstrating that it could still bind GFP (B), the resin was washed, stripped (C), and reloaded with Ni2+ (D) following standard Cube protocol (see Cube Protocols & Datasheets).

Video

Video Guide - How to use MagBeads


FAQ

Can I get the datasheet for the Ni-NTA MagBeads?

What are the reasons for nonspecific binding?

Some histidine-rich proteins can also bind to nickel. But washing with NaOH after elution of your protein of interest removes unspecific bound proteins from your resin.

I want to use a high concentration of EDTA and DTT. Is it possible to use Ni-NTA from Cube Biotech?

No, it is not recommended because nickel is reduced with DTT or dissolved with EDTA. If you want to use high concentrations of EDTA and DTT you should use our INDIGO-Ni MagBeads.

How is the capacity at high flow rates?

If higher flow rates are desired we recommend using beads with bigger diameters. We offer Ni-NTA beads with mean diameters of 40µm, 100µm, and 400µm (XL).

With each size increase, the flow rates also increase due to the proportionally increasing space between the beads. However, the surface of the beads does not increase at the same speed as the diameter (square-cube-law). That results in decreasing amounts of purified protein per mL beads while increasing the bead sizes.

For 30µm of 90µm beads, we both have average purification amounts of ~80 µg protein/mL beads. With 400 µm (XL) beads (only agarose resin, not MagBeads), this decreases to ~20 µg/mL.

We recommend reading the corresponding section of the "Guide to magnetic beads" on this subject for more detailed information.

After using DTT my resin turned orange. How to regenerate it?

The DTT has probably destroyed your beads. Ni-NTA beads only have a limited DTT tolerance of about 1 mM. However, you can regenerate them to regain their functionality. Please read our detailed protocol for more information regarding this.

However, we would recommend using Ni-INDIGO products instead. They work with the same buffers and protocols as the Ni-NTA products but have a DTT tolerance of 20 mM.