Ultrafast His-Tagged Protein Purification

Xuan Luo, Arjun S. Pamidi, Zoe Gardner, Fayed Abdullah Alrashaidi, Colin L. Raston, Gregory A. Weiss

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Abstract

This article details how to use a vortex fluidic device (VFD) to accelerate protein purification via immobilized metal affinity chromatography (IMAC). Building upon a previous report of VFD-based purification, we introduce a membrane insert to simplify the purification protocol and the resin recovery step. This new platform can be adapted to different types of IMAC resins and purification membranes. Proteins can be purified directly from clarified lysate, non-clarified lysate, and even non-lysed cultures without concerns of system clogging. Strong binding between the Ni2+ and the target protein's His6-tag effectively captures the target protein on IMAC resins or membranes placed in the VFD. Continuous flow of different solutions through the VFD allows dynamic binding, washing, and elution of the target protein. Furthermore, the system dramatically accelerates protein purification; a typical purification from cell lysate requires approximately 4 min. Herein, we demonstrate the single-step purification of two His6-tagged proteins from both clarified and non-clarified cell lysates without requiring batch binding.

Original languageEnglish
Article numbere70006
Number of pages22
JournalCurrent Protocols
Volume4
Issue number9
DOIs
Publication statusPublished - Sept 2024

Keywords

  • eGFP
  • immobilized metal affinity chromatography (IMAC)
  • membrane purification
  • protein purification
  • vortex fluidic device (VFD)

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