I want to obtain a 10% concentrated fibroin solution, but after the dialysis step, the concentration becomes between 3-5%. How can I raise it?
You could use ultrafiltration to concentrate it if it isn't viscous. Centrifugal ultrafiltration concentrator units make this very easy.
An alternative method is to draw out some of the liquid from the dialysis tubing or cassette before taking the sample out by covering it with an absorbent resin, such as Sephadex or Bio-Gel or carboxymethylcellulose.
Thank you so much Adam B Shapiro
Can I decrease it's water content by using evaporation in 50 celsius in an oven without denaturing the fibroin protein?
Dear all, why not extracting or precipitating this 3-5% and after that prepare the desired concentration. I am aware by the structural changes, but if they don't influence on the end use, there will be no problem. My Regards
Abdelkader BOUAZIZ
thanks for your response
Did you try that? What kind of structural changes occur? Does it denature?
No practical experience except theoritical knowledge. Possibly yes, denaturation occurs. My Regards
The following RG thread may be helpfull. My Regards
https://www.researchgate.net/post/Silk_fibroin_Extraction_tricks2
Evaporation by heating is certainly one way to reduce the volume, as long as it does not denature the protein. It will also concentrate any other solutes that are with the protein if it was not dialyzed against water.
An alternative to heating is freeze-drying (lyophilization). This may also cause protein denaturation, due to dehydration, and concentration of any nonvolatile solutes.
Few options we have tried,
1. Lyophiliziing small aliquots of 1% (w/v) silk fibroin solution (0.5 ml/ well) in 24 well plates and lyophilized sponges can be reconstituted to 10-12 % (w/v). The sponges can be stored in room temperature (25 deg C, desicated condition) for 2- 3 months.
2. Alternatively, 5% silk solution can be concentrated using PEG dialysis. Protocol could be found here. https://www.nature.com/articles/nprot.2011.379 (Step 22).
Joseph Christakiran Moses
Thanks,
What solvent do you use for dissolving the lyophilized Fibroin? Does it require a particular temperature for dissolving?
I have tried to resolve the lyophilized Fibroin in PBS, and also in distilled water (10% w/v) at about 50 Celsius, but it didn't completely solve.
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