Vivapure^® S Ion Exchange Membrane Adsorbers for the Rapid Fractionation of Peroxidases from Plant Calluses – Application Note

Ion exchange chromatography is one of the most-used techniques in protein biochemistry. However, as a tool for multiple sample analysis, there has been a lack of commercially available devices which offer necessary speed and performance. Recently, our laboratory became interested in a 40 kDa basic peroxidase from grapevine callus, which plays a role in rapid cell wall defense against pathogen ingress. Detailed study of this peroxidase has been limited due to its low yield from callus cultures, from which only minor quantities could be purified. 

We are looking to identify callus cultures that secrete higher levels of this peroxidase and could therefore be used as a more convenient tissue source for the purification of this molecule on a larger scale. Therefore, we intend to use cation exchange chromatography to isolate this peroxidase from other cell wall peroxidases, and subsequently quantify its abundance in different callus cultures by SDS-PAGE and measurements of peroxidase activity.

Our initial assay of different grapevine material utilized conventional SP-Sepharose column chromatography. However, we found this technique to be time-consuming and very few samples could be processed per day. It is also not amenable for fractionation of low-volume samples, which we have instead attempted to process by batch purification with sulphonic acid-based resins. In this case, high recovery of target proteins required extensive washing of the resin, resulting in sample dilution.

This application note was created in cooperation with Plant Biochemistry, ITQB NOVA, Oeiras.

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