Does anyone know whether a method exists where chloroform can be added to a recombinant protein sample after performing endotoxin clean-up to remove residual detergents? Our lab lacks access to a speedvac and we don't have the time for long dialysis processes for endotoxin removal with solvents. We attempted to use columns but we still need an initial endotoxin removal step to avoid overloading the column binding capacity. We've tried phase separation with Triton X-114 which seems to be effective but now face the problem of removing traces of the Triton which can mess with the LAL assay. I've seen protocols that use biobeads or activated carbon but we don't have these and wondered if chloroform could be used to precipitate the Triton. We attempted this and spun out the cloroform but based on a dot blot seemed to have lost the protein from the supernatant.
Does anyone know of a way to easily remove Triton, or explain any interaction and what happened to the protein?
Assuming your protein doesn't require detergent to stay in solution, I'd remove it with macro-porous hydrophobic beads like BioBeads SM2 or Amberlite XAD (see doi:10.1016/0003-2697(73)90436-3). There are also small, detergent-binding columns available meant for sample pre-processing, but if you want to do this more often, packing your own may be more economical.
You can try extraction of detergent with hydrophobic solvents, but you may lose protein on the water-solvent interface.
Please look at the following below attached papers which may help you in your analysis.
Thanks
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