Hi,
I have one question.
When express one protein (animal protein drug ) in E.coli system, E.coli endotoxin such as LPS will reside in the expressed protein. It is harmful to animal health.
If I want to produce the protein in large scale and use them as an animal drug, how could I remove the endotoxin as much as possible? And in a much cheaper way?
Thank you so much.
If you prepare recombinant proteins from E. coli, these will likely be too heavily contaminated with endotoxin to achieve efficient removal with polymixin-based endotoxin affinity resin, which will be saturated. It is better to bind your protein to a resin, like Ni-NTA if your protein bears a His-tag, and the wash extensively with ice-cold buffer containing 0.1 % Triton X-114 (the buffer has to be ice-cold, or else the detergetn will form a separate phase). Triton X-114 is then eliminated by extensive washing with detergent-free buffer (monitor A280), followed by elution with endotoxin-free buffer containing 250 mM imidazole. This will remove most of the endotoxin and the remainder can the be further eliminated by passing over the polymixin-based affinity resin. See Reichelt P, Schwarz C, Donzeau M. 2006. Prot. Expr. Purif. 46,483-488.
You can see this link
http://www.americanpharmaceuticalreview.com/Featured-Articles/190810-Removal-of-Endotoxin-from-Protein-in-Pharmaceutical-Processes/
See this link
https://www.ncbi.nlm.nih.gov/m/pubmed/10656326/
Regards
Hello,
Endotoxin can be removed with three possible mechanisms:
1. Affinity based : Affi-Prep Polymyxin resin from Bio-rad
http://www.bio-rad.com/en-in/product/affi-prep-polymyxin-resin?ID=5c53a1f3-b0da-4799-8a54-7e94f090e0f9
2. Charge based : MacroPrep High Q ( When your Protein have basic pI and operating at neutral pH) , You can try Ceramic Hydroxy Apatite also.
Selecting right resin for endotoxin removal :
http://www.bio-rad.com/webroot/web/pdf/lsr/literature/Bulletin_6813.pdf
3. Size based : If your desired protein size less then 5kda then you can try TFF (10kda) in flowthrough (permeate mode), Endotoxin have higher size so endotoxin will retain in retenate and your product will come in flow-through.
Hope this is useful.
Please contact me for any more info or support.
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