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ForteBio Interactions Newsletter Biosensor photo

SPRING 2012    VOLUME 5    ISSUE 1

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NEW  Ni-NTA Biosensors for HIS-tagged Biomolecule Analysis

The robustness of the HIS tag and its ease of use have established it as one of the most commonly used tags in the biopharmaceutical industry. It’s commonly fused to recombinant proteins to expedite their capture, detection, and purification. This polyhistidine tag sequence exhibits strong binding to nickel (Ni2+).

Pre-immobilized with nickel-charged tris-nitrilotriacetic (Tris-NTA) groups, ForteBio’s new Ni-NTA Biosensor provides easy capture of HIS-fusion proteins for kinetic analysis (FIGURE 1, TABLE 1) and rapid quantitation of HIS-tagged biomolecules (FIGURE 2). The surface offers an excellent alternative to chemical protocols such as EDC/NHS and biotinylation.

This new biosensor is fully compatible with both our Octet® and BLItz platforms. When using the Ni-NTA Biosensor with BLItz, measurement of precious samples in volumes as low as 4 µL can be determined.

The Ni-NTA Biosensor offers researchers unmatched ease of use and time-to-result in a wide range of laboratory applications including quantitation of HIS fusion proteins, affinity characterization of interactions between HIS fusions and binding partners, efficient workflows for epitope binning/mapping and process optimization in development and quality control.

For more information, please contact your local ForteBio® representative or visit our NTA biosensor page to download the datasheet and technical notes.

Kinetic analysis of the interaction between HIS-endostatin and an interaction partner anti-endostatin

FIGURE 1: Kinetic analysis of the interaction between HIS-endostatin and an interaction partner anti-endostatin. 1X Kinetics Buffer was used as the matrix throughout and the assay temperature was 30°C. Data were processed and curve fit using a 1:1 binding model. The kinetic results are reported in Table 1

 

kon koff KD
8.1X104 1/M s 1.8x10-5 1s 0.22 nM

TABLE 1: Kinetic results for the interaction between HIS-endostatin and anti-endostatin using Ni-NTA biosensors

Kinetic analysis of the interaction between HIS-endostatin and an interaction partner anti-endostatin

Kinetic analysis of the interaction between HIS-endostatin and an interaction partner anti-endostatin

FIGURE 2: Detection of a HIS-Protein A standard using Ni-NTA biosensors on an Octet RED384 system with assay parameters for a standard dynamic range. A) Assay runs at 1000 rpm and 2 minutes read time. B) Calibration curve derived from A. Sample Diluent was used as a matrix for all samples.