Hey, I work in the department that makes these, it's a whole Centrifugals department, so you'd get plenty of practice in pronunciation 😄 These are all made in Cork, Ireland, was looking for a video to show the team the end use of the devices so was delighted to find this. Keep sciencing, and sharing the knowledge.
Thanks for the wonderful video! If I wanted the final volume to be 100-200 uL using the 50 mL tube size, then some part of the membrane would become dry during the centrifuging, would this cause my protein precipitate on the membrane? What would be a better way to do this? :)
Glad you found it helpful! The protein should actually all concentrate at the bottom - it should get pulled down and therefore not get stuck on a dry membrane (but you still do have to worry about protein sticking to the membrane in general)
This video is so useful for me right now. Thank you so much. I wanna use them multiple times too, how concentrated is the ethanol you use for storage? How do you remove it before using it again?
They're typically regenerated cellulose chosen because they're hydrophilic and highly compatible with lots of different things and have low protein binding www.sigmaaldrich.com/US/en/technical-documents/technical-article/protein-biology/protein-concentration-and-buffer-exchange/protein-sample-ultrafiltration
Im using 100kda filter and my protein of interest is 40kda, can I still concentrate using that size filter? And where my protein of interest located after filtering in the 1.5 ml tube or inside the filter? Thank you
@@thebumblingbiochemist do you mean that we can push out the small peptide out of the membrane to the collection tube. Whether i can use this technique to seperate out peptides from the larger protein..thanks for ur reply
Regarding using Amicon 100 KDa, etc. I found a protocol using 3000 g for 5 minutes. My centrifuge can do only 1400 g. Is it possible to do ultrafiltration in this conditions ? You have a practice experience.
I wonder how well these will do with lipid nanoparticles. . . really fine ones. Like 30 nm in size... tried super fast centrifuging like 50k Gs for 30 minutes... didn't work so well.
@@thebumblingbiochemist thanks. I though 50 kda would would looks like I miss read. -30 would be better. My particles reach 11-12nm at the minimum. So very tiny...
Thank you for this video, was very well explained and easy to understand. I wanted to ask you, do I need to calculate my protein concentration by an assay (for ex. Bradford Assay) before using the centrifugal filter?
Hi. May I know how many percent of Ethanol are you using to store the centrifugal tubes? And What temperature you store it? For example using the Amicon ultra 15, how many mL will you pour the Ethanol?
Hey thank you for your video! It was very helpful. I am kinda new at this and I am lost with the preparation of the tubes with resin and buffer. Any resources or tips about it? Is it needed even for concentration?
This is cool! I'm doing purification and buffer exchange so this is helpful. I appreciate the details in the description and the images on the site
Thanks - glad it was helpful! And best of luck with the experiment
This has been amazingly helpful!! You're a star 🌟
Glad it was helpful!
Everytime I search for some lab info at TH-cam I end up in your channel XD would love to have you as a lab mate, tysm for sharing your knowledge!!
Happy to help!
Hey, I work in the department that makes these, it's a whole Centrifugals department, so you'd get plenty of practice in pronunciation 😄
These are all made in Cork, Ireland, was looking for a video to show the team the end use of the devices so was delighted to find this.
Keep sciencing, and sharing the knowledge.
oh awesome! thank you!
I'm purifying some protein as well and your vids have become so helpful. Thank you
So glad to hear! Good luck!
Thanks for the wonderful video! If I wanted the final volume to be 100-200 uL using the 50 mL tube size, then some part of the membrane would become dry during the centrifuging, would this cause my protein precipitate on the membrane? What would be a better way to do this? :)
Glad you found it helpful! The protein should actually all concentrate at the bottom - it should get pulled down and therefore not get stuck on a dry membrane (but you still do have to worry about protein sticking to the membrane in general)
This video is so useful for me right now. Thank you so much. I wanna use them multiple times too, how concentrated is the ethanol you use for storage? How do you remove it before using it again?
Glad to hear it! I don't remember exactly what I stored it in, but if you do a lot of buffer washes it should be good to go. Good luck!
Hlo ... Can u plz tell by which material this memberane is made up of and why only that material is used as a memb instead of any other one ???
They're typically regenerated cellulose chosen because they're hydrophilic and highly compatible with lots of different things and have low protein binding www.sigmaaldrich.com/US/en/technical-documents/technical-article/protein-biology/protein-concentration-and-buffer-exchange/protein-sample-ultrafiltration
Im using 100kda filter and my protein of interest is 40kda, can I still concentrate using that size filter?
And where my protein of interest located after filtering in the 1.5 ml tube or inside the filter?
Thank you
You need to use a much lower MWCO - something 20 or less. Otherwise, most will go through. Good luck!
Hi can we get small molecular weight peptides in the collection tube
You would need to get ones with cutoffs smaller than your peptides. Good luck!
@@thebumblingbiochemist do you mean that we can push out the small peptide out of the membrane to the collection tube. Whether i can use this technique to seperate out peptides from the larger protein..thanks for ur reply
I don't know how reliable that would be sorry
Regarding using Amicon 100 KDa, etc. I found a protocol using 3000 g for 5 minutes. My centrifuge can do only 1400 g. Is it possible to do ultrafiltration in this conditions ? You have a practice experience.
It will probably just take longer
@@thebumblingbiochemist m
Many thanks.
I wonder how well these will do with lipid nanoparticles. . .
really fine ones. Like 30 nm in size... tried super fast centrifuging like 50k Gs for 30 minutes... didn't work so well.
never tried but maybe www.sigmaaldrich.com/US/en/technical-documents/technical-article/analytical-chemistry/filtration/nanoparticle-ultrafiltration
@@thebumblingbiochemist thanks. I though 50 kda would would looks like I miss read.
-30 would be better. My particles reach 11-12nm at the minimum. So very tiny...
good luck!
Thank you for this video, was very well explained and easy to understand. I wanted to ask you, do I need to calculate my protein concentration by an assay (for ex. Bradford Assay) before using the centrifugal filter?
Only if you're curious and/or if you're aiming for a specific concentration. And thanks!
Hi, can i ask whether you can do overnight dialysis at 4 degree celcius with the protein concentrator tube?
These tubes aren't designed for dialysis - instead you'd want to use a dialysis pouch
Hi. May I know how many percent of Ethanol are you using to store the centrifugal tubes? And What temperature you store it? For example using the Amicon ultra 15, how many mL will you pour the Ethanol?
20% ethanol, enough to make sure the whole membrane stays wet, and 4°C
Thank You for the video. We usually store it in 20% Ethanol.
This was SO helpful! thank you very much!
Hey thank you for your video! It was very helpful. I am kinda new at this and I am lost with the preparation of the tubes with resin and buffer. Any resources or tips about it? Is it needed even for concentration?
Glad I could help! You shouldn't have any resin when you're concentrating! Are you thinking of chromatography?
@@thebumblingbiochemist no I'm not! Okay great. I thought that would be the case from what I've read but I was not totally sure. Thank you so much :D
Thank you for the information.... great video 👍
thank you!
Hi, If I wash with MeOH 100%, can I use again with other sample?
I've used ethanol in the past - best to use with the same protein if possible
Thanks!