Go Back   Science Forums Biology Forum Molecular Biology Forum Physics Chemistry Forum > Molecular Research Topics Forum > Basic Lab Protocols and Techniques
Register Search Today's Posts Mark Forums Read


Protein precipitation and overdried proteins

Protein precipitation and overdried proteins - Basic Lab Protocols and Techniques

Protein precipitation and overdried proteins -


Reply
 
LinkBack Thread Tools Display Modes
  #1  
Old 04-25-2008, 07:18 AM
Pipette Filler
Points: 2,584, Level: 32 Points: 2,584, Level: 32 Points: 2,584, Level: 32
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: Aug 2006
Posts: 20
Thanks: 0
Thanked 1 Time in 1 Post
Default Protein precipitation and overdried proteins



Hello,

I have a question about protein precipitation. I precipitate my proteins with methanol and chloroform. The final step is that I wash my pellets with methanol (1ml)and centrifuge down. Because of the fragility of the pellet I put it in all in the speed vac and dry the pellet. What could happen with the proteins when I overdry the pellet. After drying I resuspend the pellet with 2x sample buffer. When I have loaded the protein on the SDS-PAGE I observed that in the bottom of the tube a residue is visible. Something viscous or the like. Is this the overdried pellet and the proof that not everything was properly resuspended?
Thanks for all comments.
Tiffy
Reply With Quote
  #2  
Old 04-25-2008, 02:50 PM
danfive's Avatar
Nobel Laureate
Points: 8,612, Level: 64 Points: 8,612, Level: 64 Points: 8,612, Level: 64
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: Jul 2007
Location: Houston TX
Posts: 969
Thanks: 12
Thanked 184 Times in 154 Posts
Default Re: Protein precipitation and overdried proteins

In best case scenario the overdried pellet can still be resuspended by adding time, sonication, gentle grinding, or using a stronger resuspension buffer. Worst case scenario you'll lose the overdried proteins and they won't resuspend.

Sometimes you need to have complimentary protein extraction/isolation methods for proteins based on their chemical characteristics: hydrophobic/hydrophilic proteins, glycoproteins, large mass proteins, high abundance/low abundance etc.

There isn't a perfect method for isolating 100% of proteins, you have to tailor your extraction/isolation to suit your target proteins.
Reply With Quote
Reply

Tags
overdried , precipitation , protein , proteins


Thread Tools
Display Modes

Posting Rules
You may not post new threads
You may not post replies
You may not post attachments
You may not edit your posts

BB code is On
Smilies are On
[IMG] code is On
HTML code is Off
Trackbacks are On
Pingbacks are On
Refbacks are On

Forum Jump


All times are GMT. The time now is 09:26 AM.


Powered by vBulletin® Version 3.8.4
Copyright ©2000 - 2014, Jelsoft Enterprises Ltd.
Copyright 2005 - 2012 Molecular Station | All Rights Reserved
Page generated in 0.11493 seconds with 15 queries