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How Parallel Protein Purification with Improved Yield and Concentration Works

Protein affinity purification is typically carried out in a miniaturized format using spin columns or gravity flow methods. Both techniques have limitations.

by Rainin
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Problem: Protein affinity purification is typically carried out in a miniaturized format using spin columns or gravity flow methods. Both techniques have limitations. Spin columns do not maximize the capacity of a given resin because the protein-containing solution passes through the resin bed too quickly. Due to this, the final protein yield is usually lower when compared with gravity flow or batch purification methods. The user of this technique might find that obtaining sufficient protein requires repeating the same protocol over and over.

Small-scale gravity flow protein purification better maximizes the capacity of a given resin due to the slow flow-rate of purification media over the resin bed. Gravity flow methods, however, are cumbersome because in order to maintain target protein activity, constant oversight is needed to ensure that the resin remains wet. Finally, the elution volumes for gravity flow columns are typically large, which dilutes the purified protein, and may compel the user to add an extra concentration step to obtain a sample of high enough concentration for their functional assay.

Solution: One product that may help is the PureSpeed Protein Purification System from Rainin Instrument, a subsidiary of Mettler Toledo. The PureSpeed system utilizes a resin bed in a pipette tip alongside a multichannel E4 XLS electronic pipette for parallel purification of up to 12 protein samples. This system is advantageous compared to spin column and gravity flow formats because it uses repetitive, automated up-and-down pipetting and a tightly packed resin volume to maximize protein binding to resin while reducing hands-on time and the required elution volume. The final protein yield is higher than that obtained for spin column purification while the final protein concentration is higher than that obtained for gravity flow methods. The result is the user will not have to repeat the same purification and will not have to carry out downstream concentration steps prior to assaying the protein sample. Finally, compared to gravity flow methods, the user does not have to worry about protein inactivation due to the resin drying.

The PureSpeed system has many accessories and features available to the user so they can easily customize and carry out their processes. For example, Rainin PureSpeed protein tips are available with three different types of resin: protein A and protein G resins are used to purify antibodies, while Ni-IMAC resin is used to purify 6 x His tagged proteins. With the E4 XLS pipette and PureSpeed Accessory Kit, PureSpeed tips can be used for semi-automated up-and-down pipetting in protein loading, washing, and eluting steps. The user, after setting up the PureSpeed system with the appropriate buffers and protein samples in a deepwell plate, can initiate the PureSpeed protocol on the E4 XLS pipette prior to stepping away from the purification system until the next step in the procedure. Following each step, the pipette is simply transferred from one row of wells in the deepwell plate to the next, and after completion of the protocol, purified protein is easily collected from the elution wells. The final protein yields and concentrations are highly reproducible using PureSpeed mode with the E4 XLS pipette. In addition, protein functionality is preserved throughout the purification process, making many downstream assays a reality.

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The Rainin PureSpeed Protein Purification System: for automated parallel processing to obtain up to 12 protein samples of excellent purity, concentration, and yield.