Bio-Rad Profinia™ Protein Purification Instrument - Manual (Page 129)

Figure 10.2 shows proper placement of denaturing IMAC buffers in the Profinia instrument.
Fig. 10.2. Denaturing IMAC buffers properly positioned with instrument lid closed.
10.4 GST Purification and Buffer Kits
The lysis and wash buffers contained in the GST kit are formulated from sodium salts,
phosphate buffers, and EDTA, and provide optimum lysing, binding, washing, and elution
of GST-tagged proteins. EDTA is included as a chelating compound and protects against
metalloproteases, a class of proteases that can be present in bacterial lysates. GST fusion
proteins must be enzymatically active prior to purification. The buffers used in this system
are designed for the purification of proteins that partition into the soluble fraction of bacterial
lysates and that have accessible, and biologically active, GST sequence tags. Table 10.4
provides a list of buffer compositions. The lysis buffer is used for sample preparation (see
Appendix E), and is not used in any of the buffer bottle positions.
The GST cartridges are provided in 20% ethanol, and should be stored at 4°C prior to use.
The cartridges are packed with 1 ml of Bio-Rad Profinity
™ GST resin, and typically yield ~10
mg of purified protein per run (dependent upon protein expression level and culture volume
loaded). To minimize any possibility of cross-contamination, it is suggested that individual
cartridges be dedicated to the purification of a unique GST-tagged protein. The desalting
cartridges are provided in a PBS/0.05% azide buffer and can be stored at 4–22°C prior to
use.
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