I induced a protein with halo tag. And when run my sample( still on the beads) as well as lysis, I found that my protein has a very limitted amount binding to the beads
Hi there,
THere might be several issues. Have a read at the troubleshooting section of the Promega leaflet below (p.24):
https://www.promega.com/-/media/files/resources/protocols/technical-manuals/0/halotag-protein-purification-system.pdf
thanks very much for your help. I do read it. And it doesn’t help me
old thread. If you are still following it, more detail is needed. I just tried capture of a Halo fusion at 3 uM fusion in E coli lysate with 5:1 ratio of lysate to Promega's HaloLink resin, and got 75% capture in 1 hr at room temp. This is the level of detail you need to provide to get help you debug. I also found that Halo was resistant to 50% urea -- still can be labeled with halo ligand efficiently (although I did not look at kinetics with/without urea side by side).
Andrey Revyakin I have background issues with promega halo beads in mammalian cells. Do you have any suggestions how to reduce it? Could you please share the protein and bead amount you used for IP. The promega leaflet wasn't helpful in this context.
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