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Immobilization of antibodies

Immobilization to NHS beads

Antibodies can be immobilized on NHS beads. NHS beads are beads where the COOH groups on the surface have been activated by succinimide. A reaction is produced easily by mixing these beads with a substance that contains NH2 groups in order to create antibody-binding beads, without any additives or special pretreatment. When antibodies are mixed with the NHS beads, the ε-NH2 groups of the antibody lysine residue form amide bonds with the activated COOH groups on the surface of the FG beads®. If NHS groups which have not bonded with antibodies are present, they separate in the solution and the COOH groups are exposed to the surface, causing the surface of the beads to become charged and resulting in electrostatic protein adsorption. It is therefore necessary to use aminoethanol to perform masking of the remaining NHS groups.

Because the antibodies and FG beads® are bound by covalent bonds, it is possible to prevent antibody elution when the target proteins are eluted with acid.

Immobilization to Epoxy beads

Antibodies can be immobilized on epoxy beads. The ε-NH2 groups of the antibody lysine residue or the imidazole groups of the histidine residue form covalent bonds with the epoxy groups on the bead surfaces. The FG beads® and antibodies form covalent bonds, preventing the antibodies from being eluted when the target proteins are eluted using acid. Because epoxy beads produce almost no non-specific protein adsorption, it is not necessary to mask the remaining epoxy groups.

No.Protocol
105Immobilization of Antibodies or Proteins on NHS Beads
106Immobilization of Antibodies or Proteins on Epoxy Beads

Immobilization to streptavidin beads

Biotin-modified antibodies can be bound to streptavidin beads. When immunoprecipitation is performed using streptavidin beads with biotin-labeled antibodies bound to them, it is possible to achieve a higher level of antigen purity compared with the streptavidin beads of other companies.

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