Good afternoon. I am writing a scientific paper on the production of recombinant TNF-alpha. I am focusing on the downstream process. The steps are two-step ion exchange chromatography. And then there is a desalting step with dialysis. But I don't know what kind of membrane to use and what pore size to use. TNF-alpha is 17 KD. I would also like to know if dialysis can be scaled up in industry?
Thank you for your Answer.
I think you could use 10K MWCO dialysis tubing/membrane for your 17 kDa protein. This Thermo Fisher note is quite helpful for understanding the relationship between the protein's molecular weight and the cutoff size of the dialysis membrane.
https://www.thermofisher.com/mx/es/home/life-science/protein-biology/protein-biology-learning-center/protein-biology-resource-library/protein-biology-application-notes/separation-characteristics-dialysis-membranes.html
use a membrane with a molecular weight cut-off (MWCO) of 10-15 kDa to retain the protein. Opt for low-protein-binding materials like regenerated cellulose or polyethersulfone (PES) to minimize loss. Ensure compatibility with the buffer system and maintain gentle handling to preserve protein integrity.
You can use a membrane with 10kDa or lower cutoff (the cutoff value should be ~2x lower than the protein size, sometimes you can get surprises with larger protein going into or through the membrane pores if the difference is too small).
As an alternative (and better for sacle-up), TFF with buffer exchange and concentration of the sample is more convenient (for smaller scale you could try the Vivaflow cartridges from Sartorius).
Or you could use Sephadex™ G-25 resin for faster buffer exchange (like PD10 column) which will dilute the sample a bit. The resin is not too expensive and can be used with larger volumes using empty columns in different sizes.