StrepTactin is a specially engineered streptavidin ligand. The binding afﬁnity of Strep-tag II to the immobilized ligand is nearly 100-fold higher than to streptavidin, making StrepTactin Sepharose® High Performance ideal for purifying Strep-tag II proteins.
The small bead size (average 34 µm) of the Sepharose® High Performance matrix results in high-resolution separations, sharp peaks, and puriﬁed target proteins in a concentrated form. StrepTactin Sepharose® High Performance is compatible with a wide range of additives, tolerates all commonly used aqueous buffers, and is quickly and easily regenerated using 0.5 M NaOH. See Appendix 4 (Characteristics of StrepTactin Sepharose® High Performance products) for more information on the characteristics of the medium.
See instructions supplied with the product or refer to Appendix 6 (Column packing and preparation) for general guidelines for column packing.
Adjust the sample to the composition of the binding buffer (see below). For example, dilute the sample with binding buffer or buffer exchange using a desalting column see Chapter 11 (Desalting/buffer exchange and concentration).
Pass the sample through a 0.22 µm or a 0.45 µm ﬁlter and/or centrifuge it immediately before applying it to the column. If the sample is too viscous, to prevent it from clogging the column dilute it with binding buffer, increase lysis treatment (sonication, homogenization), or add DNase/RNase to reduce the size of nucleic acid fragments.
See Appendix 4 (Characteristics of StrepTactin Sepharose® High Performance products) for details on regeneration of the medium.
Water and chemicals used for buffer preparation should be of high purity. Filter buffers through a 0.22 µm or a 0.45 µm ﬁlter before use.
The recommended ﬂow velocity is 150 cm/h.
Scale-up is typically performed by keeping bed height and ﬂow velocity (cm/h) constant while increasing bed diameter and volumetric ﬂow rate (mL/min). See Figure 7.4 (Puriﬁcation using StrepTrap HP 1 mL and 5 mL) for an example of scale-up using this medium.
Store StrepTactin Sepharose® High Performance in 20% ethanol at 2 °C to 8 °C. After storage, equilibrate with binding buffer before use.
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