Our Halo-Trap is a high affinity Halo-binding protein coupled to agarose beads for biochemical analysis of Halo fusion proteins and their interacting partners. The Halo is a modified variant of the bacterial haloalkane dehalogenase enzyme from Rhodococcus rhodochrous and is designed to form a covalent bond with reactive chloralkane-based ligands.
Use Halo-Trap for:
- Immunoprecipitation (IP) / Co-IP
- Mass spectrometry
- Enzyme activity measurements
- ChIP/ RIP analysis
Very stable & reliable binding
- high affinity with a very low dissociation constant KD of 2nM
- harsh washing conditions can be applied
- recombinant, validated antibody
- Structure and function are characterized
Halo-protein immunoprecipitated with either Halo-Trap (left) or an alternative supplier's product (right): Recombinantly expressed Halo-protein bound to Halo-TMR. Input (I), non-bound (FT) and bound (B) fractions were separated by SDS-PAGE followed by Coomassie blue staining, Western blotting and fluorescence scan. Please note, additional bands are visible due to sensitivity of the Halo-protein to SDS. Because the Halo-Trap binds to Halo-tag at an epitope outside its catalytic center, Halo-proteins covalently bound to ligands like Halo-TMR (left) are effectively captured. In contrast, the competitor’s product precipitates no or just minimal amounts of the Halo protein. This allows to pulldown Halo-fusion proteins from cells that have been previously treated with Halo-ligands for imaging and makes the Halo system as versatile as GFP expression, imaging and purification.
Compared to other Halo tools,
- Halo-Trap precipitates Halo-fusion proteins even when already covalently bound to Halo-ligands, i.e. after labelling etc.
- Halo-Trap can be used for affinity purification
- bound Halo-fusion protein can be eluted without protease
Immunoprecipitation and elution of Halo-protein from spiked HEK293T cell lysates: Input (I), non-bound (FT), eluted (E), residual (R) fractions and protein marker (M) were separated by SDS-PAGE followed by Coomassie blue staining and Western blotting. Elution of Halo-protein with elution buffer (low pH) from Halo-Trap is efficient (left).