Tetanus toxoid (TT) is frequently used as a carrier protein for conjugate vaccines. TT contains strong T-cell epitopes. However, TT is a toxoided protein and is not a uniform product because of specific toxoiding and purification processes. Full-length TT tends to aggregate with age and is not affordable to produce in quantities for research and early clinical work. In contrast, TT heavy chain fragment C (rTTHc; the C terminal fragment of the toxin) is devoid of the toxin’s enzymatic activity but contains the binding site necessary for uptake of the toxin by neurons. rTTHc is also the most immunogenic part of the toxin, containing the powerful T-cell epitopes of TT without the toxicity.

In this case study, Andrew Lees, PhD presents a purification process for rTTHc using polymeric chromatography resins. rTTHc was expressed at high levels as a soluble, properly folded intracellular protein in Fina Biosolutions proprietary Escherichia coli strain. Multiple resins were screened using an automated chromatography system. An anion exchange (AEX) resin — Nuvia Q Resin — was selected as the capture step and Macro-Prep Methyl Hydrophobic Interaction Chromatography (HIC) Resin selected for the polishing step. This two-step purification protocol was optimized for scale-up, resulting in greater than 98% pure purified target protein.


  • Importance of conjugate vaccines
  • Strategies for purifying carrier proteins
  • Optimized screening methods for ion-exchange (IEX) and HIC resins

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