Tetanus toxoid (TT) is frequently used as a carrier protein for conjugate vaccines. TT contains strong T cell epitopes. They are toxoided protein and are not a uniform product because of specific toxoiding and purification processes. TT tends to aggregate with age and is not affordable to produce in quantities for research and early clinical work. TT heavy chain fragment C (rTTHc) is the C terminal fragment of the toxin. rTTHc is devoid of the toxin’s enzymatic activity but contains the binding site necessary for uptake of the toxin by neurons and is the most immunogenic part of the toxin. Fragment C contains the powerful T cell epitopes of tetanus toxin without the toxicity.
In this case study, Dr. Lees presents the purification process for rTTHc using polymeric chromatography resins. rTTHc was expressed at high levels as a soluble, properly folded intracellular protein in Fina Bio proprietary Escherichia coli strain. Multiple resins were screened using an automated chromatography system. Nuvia Q anion-exchange (AEX) resin (120 µm) was selected as the capture step and Macro-Prep Methyl hydrophobic-interaction chromatography (HIC) resin selected for polishing step. This two-step purification protocol was optimized for scale-up, resulting in greater than 98% pure purified target protein.
Overview
Importance of conjugate vaccines
Strategies for purifying carrier proteins
Optimized screening methods for ion-exchange (IEX) and HIC resins
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Негізгі бет Efficient and Rapid Purification of E. coli Expressed Toxin Recombinant Protein Fragments
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