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

SDS-PAGE Gel Electrophoresis Forum SDS-PAGE Forum and Gel Electrophoresis Forum. Discuss the running of agarose gels, sds-page gels, and other gels including sequencing, gradient or tricine.


blurred bands after silver staining

blurred bands after silver staining - SDS-PAGE Gel Electrophoresis Forum

blurred bands after silver staining - SDS-PAGE Forum and Gel Electrophoresis Forum. Discuss the running of agarose gels, sds-page gels, and other gels including sequencing, gradient or tricine.


Reply
 
LinkBack Thread Tools Display Modes
  #1  
Old 06-23-2011, 06:11 AM
Pipette Filler
Points: 816, Level: 16 Points: 816, Level: 16 Points: 816, Level: 16
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: Jun 2011
Posts: 4
Thanks: 0
Thanked 0 Times in 0 Posts
Default blurred bands after silver staining



hi....
can any one help me. I am using 10% gel. and running at 100v and i am using the bio-rad apparatus. after silver staining the bands are not clear the whole lane in which sample was run is taking the stain and the bands are also not clearly resolved. i have tried running the experiment a number of times but the problem persists. the sample is a plant sample and protein has been extracted via tris-cl extraction method and desalted using bio-rad biospin 6 bio-gel column followed by lypholization and reconstituting in 65 micro litre 5X sample buffer and adding to it 4 micro litre beta mercaptoethanol as the total loading volume supported is 70 micro litre

the stock solution of sample buffer does not contain beta mercaptoethanol we add it later
i am uploading the image kindly tell me what should i do. we dont know anything about the protein as no work has been done on it before
Attached Images
File Type: jpg 23062011330.jpg (90.8 KB, 9 views)
Reply With Quote
  #2  
Old 08-06-2011, 04:26 AM
Volunteer
Points: 231, Level: 4 Points: 231, Level: 4 Points: 231, Level: 4
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: Aug 2011
Location: Malaysia
Posts: 36
Thanks: 0
Thanked 14 Times in 13 Posts
Default Re: blurred bands after silver staining

its looks like you've overloaded your protein samples. I have experience with plant proteins and i do face extraction problems initially with plant proteins compared to other organisms. Maybe you can try changing your protein extraction protocol, e.g. phenol extraction/ TCA-acetone extraction/ buffer with sucrose...etc. you can try separating your proteins on 12.5% resolving & 5% stacking SDS-PAGE, with lower concentration of your proteins, maybe 2ug.
Reply With Quote
The Following User Says Thank You to JennYeap For This Useful Post:
admin (08-06-2011)
Reply

Tags
bands , blurred , blurry , problem , silver , silver staining , staining


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
Problem with Sweet silver staining of proteins Lenu Molecular Biology Techniques 0 06-02-2011 10:59 AM
Immidiate dissappearing of bands on DNA sequencing gels after silver staining himanshu SDS-PAGE Gel Electrophoresis Forum 2 07-24-2008 06:17 PM
Problems with silver staining terence diane fabella Protein Forum 2 01-16-2008 06:04 PM
USP silver staining method help leelavati shetty Protein Forum 0 01-16-2008 05:17 AM
Silver staining Western membranes EK Protocols and Methods Forum 14 09-27-2003 02:37 PM


All times are GMT. The time now is 05:25 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 0.12653 seconds with 17 queries