The AminoLink Immobilization Kit contains AminoLink Coupling Resin and associated reagents sufficient to perform 5 protein immobilizations, with expected an expected yield of 2 to 20 mg protein per column. AminoLink Coupling Resin is crosslinked 4% beaded agarose that has been activated with aldehyde groups to enable covalent immobilization of antibodies and other proteins through primary amines.
Features of AminoLink Coupling Resin:
• AminoLink Coupling Resin—aldehyde-activated crosslinked, 4% beaded agarose • Ideal for antibodies and other proteins—immobilize molecules via primary amines (-NH2) • Flexible coupling conditions—efficient (>85%) coupling over a wide range of pH (4-10) and buffer conditions (PBS or other non-amine buffer with or without organic solvent) • Stable, permanent immobilization—Coupling reaction results in stable, leak-resistant secondary amine bond between resin and ligand • Better than immobilization to CNBr-activated agarose—bond is more stable and uncharged, resulting in less nonspecific binding in affinity purification procedures • Versatile and reusable—prepared affinity resin is adaptable to column and batch affinity techniques and the resin is reusable for typical applications based on protein binding interactions
Proteins and other molecules with primary amines can be covalently attached (immobilized) to AminoLink Resin to make chromatography columns for use in affinity purification. The aldehyde groups form stable secondary amine bonds with primary amines such as exist in the side chain of lysine (K) residues, which are generally abundant and readily accessible in proteins. Once a protein is immobilized, the prepared affinity resin can be used for a variety of batch and column affinity purification methods involving binding interactions with the immobilized protein. The resin and linkage are stable in most binding and elution conditions typically used in affinity chromatography, enabling prepared resin to be used for multiple rounds of affinity purification procedures.
The AminoLink Resin immobilization reaction involves spontaneous formation of Schiff base bonds between aldehydes and amines and their subsequent stabilization by incubation with a mild reductant. The entire coupling reaction, called reductive amination, occurs in 4 to 6 hours in simple non-amine buffers such as PBS. Coupling efficiency with antibodies and typical proteins is generally greater than 85%, resulting in 1 to 20 mg of immobilized protein per milliliter of resin.