Jump to content

  • Log in with Facebook Log in with Twitter Log in with Windows Live Log In with Google      Sign In   
  • Create Account

Submit your paper to J Biol Methods today!
Photo
- - - - -

Transient Transfection


  • Please log in to reply
1 reply to this topic

#1 Gunasekaran

Gunasekaran

    member

  • Members
  • Pip
  • 2 posts
0
Neutral

Posted 15 February 2002 - 01:14 AM

Hellow!
I have a problem with transient transfection of Rat-2 fibroblast cells.
I intend to assay the luciferase-reporter gene activity.
The beta-gal is the internal control to check the transfection efficiency.
Every time I get inconsistent results although I use same reagents, same DNA .
What could be the problem?
Secondly, when I do luciferase assay, the value drastically changes between two samples with similar treatment...more funny is that the same sample, when taken reading at different time, the value changes.
By different time, I mean within 5 minutes.
What could be the problem?
Waiting for your reply.
Thanks!

#2 Peter Terry

Peter Terry

    member

  • Members
  • Pip
  • 4 posts
0
Neutral

Posted 26 February 2002 - 06:54 AM

Inconsistency of results of the tests means that the substrate you are putting into the tests is inconsistent, whether you realize it or not ie: you have a quality control problem.  There are many factors which may be varying.  I suggest you list all the aspects of the experiment (eg: cell plating density, time to transfection, size of samples for luminometer etc and define how you measure each.  You will find sources of variation.  I have found the www.lifetech.com to be a good source of assistance.  Time spent analyzing all these little problems is better than time spent repeating mistakes




Home - About - Terms of Service - Privacy - Contact Us

©1999-2013 Protocol Online, All rights reserved.