I have been trying to dissolve human fibrinogen in different solvents including 0.9% saline with/out aprotinin, 1.7% Saline, PBS, PBS+Calcium Chloride, Tris Buffer, . However, no matter what solvent I use, It takes several hours , sometimes even 1 day to get it dissolved. I normally put the solution in a 37 C degree incubator and shake it by hand moderately every 1-2 hour. I tried dissolving fresh fibrinogen concentrate obtained via chemical precipitation before lyophilization, hoping to get a shorter dissolution period but dissolution time was even longer for fresh concentrate.
I wonder if anyone here has a similar experience with fibrinogen and can advise what trick/strategy possibly can cut the time of fibrinogen dissolution shorter?
Thanks everyone in advance for valuable comments shared.
#Fibrinogen #Fibrin #Dissolution time
Must try sonication for long period of time.
At temp. around 40C, the fibrinogen dissolves very slowly at 1% saline giving a slight haziness to liquid but their will be some undissolved parts left behind. Changing the pH also is in effective.
Prashant Pandey
Hi Prashant,
Thanks for your advice but i wonder wont long time sonication destroy protein structure of fibrinogen and make it less clottable? and how "long" is expected to be enough? I mean how many mins or hours? because i already tried 2 min and it does not effect the solubility, even it seemed sonication have clumped fibrinogen particles into a more dense mass after 2 min of sonication. the fibrinogen was suspended in a 0.9% saline.
Ok, I discussed with someone about your issue.
Hey suggested that search some papers in which fibrinogen treated with Tris-HCl buffer.
@prashant pandey
Thanks for time spent. But Tris buffer is cyrotoxic to my knowledge and i want to use fibrinogen for cell seeding scaffold fabrication. Any idea avout sodium citeate suitability?
I just add PBS to reach 25 mg/ml concentration and then rock it overnight at 4OC. Never agitate it by hand
Mohamadhasan Ansarizadeh Thanks for your contribution. U mean going beyond 37 C and using an automatic shaker overnight should resolve the issue?
No, I mean 4 centigrade degrees not 40! put it on rotator in the cold room where the temperature is +4.
Hi Rooney
Did it work for you? I am attempting to purify fgn, which buffer has worked for you finally. This will give me a direction too.
Thanks
Sneha Singh well ,the best solvent indeed was plasma itself, i.e. re-suspending the concentrate in plasma to reach the desired concentration. but hepes/tris/PBS and even citrate will also work so long as to keep pH around 7 and using physical (not chemical) precipitation method to purify fibrinogen. chemically precipitated fg will solve at a much slower rate. perhaps due to binding of small ligand molecules used in chemical precipitation methods which needs to be removed before re-suspending fg in solvent.
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