Is it necessary to add lysozyme if xTractor Buffer is used with mammalian cells?

xTractor Buffer can be used with mammalian cells at efficiencies comparable to those obtained for bacterial pellets. However, since mammalian cells are easier to extract for protein, lysozyme does not need to be added to the lysis buffer.Protein purification products Care guide Caution [body] This care sheet provides general information only for handling Carolina™ bacterial cultures. When you [...]

What is xTractor Buffer?

xTractor Buffer is an exceptionally flexible and efficient lysis buffer for protein extraction from bacterial, yeast, mammalian, and baculovirus-infected cells. It provides a fast and efficient 10 min lysis using mild, nondenaturing conditions. xTractor Buffer has been optimized for extraction of his-tagged proteins, so it is compatible with all IMAC resins.Protein purification products Care [...]

Can arginine be present in buffers used with TALON resin?

Arginine may be used in buffers intended for use with the TALON resin only if the final concentration is lower than 1 mM. At higher concentrations (and if using an elevated pH for adsorption) it would likely interfere with the binding of the his-tagged protein. If using an arginine-containing buffer, the pH should be maintained at ~7.2 and the sample diluted if the final arginine concentration [...]

Should a pH- or imidazole-based elution be used?

This is the researchers' choice. We recommend using imidazole since it minimizes nonspecific binding, but imidazole impurities and his-tagged proteins both absorb at 280 nm, so elution peaks can be difficult to distinguish. However, Takara Bio's buffers are made with low-absorbance imidazole, so the only molecule absorbing at 280 nm is the his-tagged protein. The choice may also depend on [...]

Can I elute my his-tagged protein from TALON resin with a histidine gradient?

Histidine is not recommended for protein elution from TALON resin because it will bind strongly to cobalt ions and cause metal ion transfer, resulting in stripping of the column. We recommend imidazole because it lacks the additional primary amine of histidine, so it binds less strongly to the metal ions and does not cause stripping of the column.Protein purification [...]