Go Back   Science Forums Biology Forum Molecular Biology Forum Physics Chemistry Forum > Molecular Research Topics Forum > Protein Science > Western Blot Forum
Register Search Today's Posts Mark Forums Read

Western Blot Forum Discuss western blotting and immunoblot in the western blot forum discussion board. Ask Questions about transfers, blocking, membrane antibody incubation and exposures.


Electrophoresis Problems

Electrophoresis Problems - Western Blot Forum

Electrophoresis Problems - Discuss western blotting and immunoblot in the western blot forum discussion board. Ask Questions about transfers, blocking, membrane antibody incubation and exposures.


Reply
 
LinkBack Thread Tools Display Modes
  #1  
Old 03-05-2007, 09:17 AM
Pipette Filler
Points: 1,405, Level: 22 Points: 1,405, Level: 22 Points: 1,405, Level: 22
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: Mar 2007
Posts: 2
Thanks: 0
Thanked 0 Times in 0 Posts
Default Re: Western blooot troubleshooting



Hello everybody..I have elecrophorised some whole cell extracts but I only got single huge bands in each lane, while the marker has run perfectly..Could anyone help me out to solve the problem?

Thank you..
Reply With Quote
  #2  
Old 03-06-2007, 03:58 AM
moleculardude's Avatar
M.D/Ph.D
 
Join Date: Jul 2006
Location: Melbourne Australia
Posts: 423
Thanks: 5
Thanked 10 Times in 8 Posts
Default Re: Electrophoresis Problems

Hey Missmel!

Welcome to the forums!

Wow thats a weird one. I think you have a problem with your sample preparation. It may be your SDS-PAGE loading buffer that is the problem.

If your sample buffer has too much sucrose this may cause the problem.

I would remake your SDS-Sample buffer or get some off someone that does not have this problem.

Also could be your samples themselves. Maybe too much protein or salt concentration.

Please give us a bit more information and we can help you more!
please tell us the outcome you got,
Cheers
Reply With Quote
  #3  
Old 03-12-2007, 04:07 PM
Pipette Filler
Points: 1,405, Level: 22 Points: 1,405, Level: 22 Points: 1,405, Level: 22
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: Mar 2007
Posts: 2
Thanks: 0
Thanked 0 Times in 0 Posts
Smile Re: Electrophoresis Problems

Thanx for the quick response.
Now that you mention it, I use a non-reducing sample buffer (no mercaptoethanol, or DTT) 'cause the antibody detects a non-reduced epitope...
Could that be causing the problem? If yes, any suggestions as to how to bypass it?

thanx again
Reply With Quote
  #4  
Old 05-28-2007, 04:26 PM
Pipette Filler
Points: 1,284, Level: 20 Points: 1,284, Level: 20 Points: 1,284, Level: 20
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: Apr 2007
Posts: 5
Thanks: 0
Thanked 0 Times in 0 Posts
Default Re: Electrophoresis Problems

Yes, this is your problem....
you should just reduce acrylamide % and use a non reduced marker....but you must remember that when you run non denatured samples, the migration is not only dependent from molecular weight, but also from secondary and tertiary structure of the protein.
Reply With Quote
Reply

Tags
electrophoresis , problems


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

Similar Threads
Thread Thread Starter Forum Replies Last Post
Problems with PCR, gel electrophoresis, sequencing biggy1001 PCR - Polymerase Chain Reaction Forum 7 09-08-2009 09:03 PM
problems with growing healthy Arabidopsis plants Monika Kuzma Arabidopsis and Plant Biology 1 02-19-2007 03:55 PM
Problems with growing healthy Arabidopsis plants - light intesity? Karl Lundy Arabidopsis and Plant Biology 0 02-07-2007 05:11 PM
[ANN] xmds-1.3-4 released! xmds solves complex problems simply and quickly Paul Cochrane Forum Chemie 0 06-21-2004 01:29 AM


All times are GMT. The time now is 11:27 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 0.13006 seconds with 16 queries