Anti-His-tag capture system: fast, reversible and convenient protein immobilization

The anti-His-tag capture system immobilizes His-tagged proteins on an oligonucleotide sensor chip through the Oligo Adapter trisNTA. This application note tests it with human Fc gamma receptor I (FcγRI) and the therapeutic antibody cetuximab.

The Challenge: Reversible Capture of Recombinant Proteins

The His-tag is a widely used handle for recombinant proteins. It binds nickel ions held by a chelator such as nitrilotriacetic acid (NTA). For kinetic work, this capture must stay stable during the measurement, and the sensor chip surface should be regenerated afterward.

The Approach: Oligo Adapter trisNTA, FcγRI and Cetuximab

Oligo Adapter trisNTA (100 nM) was hybridized to an oligonucleotide sensor chip on a MACS Matchmaker and charged with 500 µM NiSO₄. His-tagged FcγRI (100 nM) was captured next. Single-cycle kinetics in PBS-T used cetuximab from 3.91 to 1000 nM, with a final dissociation of 120 minutes. A parallel buffer injection corrected for the decay of the capture surface.

Key Results for Cetuximab Binding to FcγRI

• Affinity: K_D of 9.0 ± 5.7 nM from a Langmuir 1:1 fit.
• Replicates: 63 molograms analyzed in a single flow chamber.
• Specific capture: FcγRI was captured through its His-tag after the adapter was loaded with Ni²⁺.
• Decay correction: the buffer control compensated for the slow loss of ligand from the capture layer.

Why It Matters for Recombinant Proteins

Many recombinant proteins already carry a His-tag, so this system needs no extra coupling chemistry. Multivalent binding between the trisNTA head and the His-tag strengthens the complex. The adapter is held by hybridization, so the sensor chip surface can be regenerated with the defined regeneration.