1. Affinity Resin

PureCube Co-NTA Agarose was developed for the affinity purification of proteins carrying a polyhistidine tag. This affinity chromatography matrix is based on BioWorks Workbeads, consisting of 7.5% crosslinked agarose. The material is highly porous to allow for optimal protein interaction. Cross-linked agarose is also physically very stable, making it suitable for purification processes under low pressure with variable flow rates. Our agarose is very homogeneous in size with a medium particle diameter of 40 µm, yielding a high degree of reproducibility between individual purification runs.

An NTA ligand is coupled to the agarose resin and carefully loaded with cobalt ions to obtain a matrix with high specificity and binding capacity for histidine residues. The metal ion capacity is about 15 µeqv Co2+/mL. Other possible metal ions are Ni2+, Zn2+, Fe3+, Al3+, resulting in different affinities, e.g., for zinc-finger proteins or phosphorylated proteins. If required, the nickel ions can be removed from the agarose matrix using five wash steps with 100 mM EDTA, and the matrix can be recharged with a different metal ion. Alternatively, please contact us for unloaded NTA agarose. NTA matrices charged with other metals are available upon request.

2. Protein Binding Capacity

PureCube Compact Cartridges Co-NTA have a binding capacity of up to 20 mg/mL as determined by purification of 6xHis-tagged GFP protein from E.coli cleared lysates, and quantified via spectrophotometry. It should be considered that the dynamic binding capacity strongly correlates with flow rate and other parameters such as protein size and buffer conditions. It is recommended to use the lowest flow rate possible to achieve highest binding capacity.

3. Compatibility

    For cleaning purposes, PureCube Co-NTA Agarose is very stable and can resist the following conditions in most situations: All commonly used aqueous buffers, pH 2-14, 100% methanol, 100% ethanol, 8 M urea, 6 M guanidinium hydrochloride, 30% (v/v) acetonitrile, and up to 10 mM DTT.

Features - Regarding Purification capabilities.

Features - Regarding Cartridge / Column itself.