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
- - - - -

What is the optimal time for transfection efficiency measuring?

transduction efficiency

  • Please log in to reply
1 reply to this topic

#1 ticovka

ticovka

    member

  • Members
  • Pip
  • 1 posts
0
Neutral

Posted 23 November 2013 - 10:01 AM

Dear Colleagues, I have a simple question: what is the optimal time for lentivirus transduction efficiency measuring (eg. by GFP)?

Transducted cells need not so much time to express the GFP protein, but why in most cases researchers measure the efficiency after 12-48-72 h., but not in 6 h., for example? Why we need to wait so long?? Mammalian cells, which was not transducted, divided approximately 4-8 times in this long time, for this reason the efficiency of transducted cells in % will be less after 72 h. then after 6 h., am I wright?


SO, what is the logic in choosing such time points?


Edited by ticovka, 23 November 2013 - 11:08 AM.


#2 2xzwei

2xzwei

    Enthusiast

  • Active Members
  • PipPipPipPipPip
  • 31 posts
1
Neutral

Posted 28 November 2013 - 03:59 AM

As always - it depends. The transduction procedure and overexpression of genes can lead to stress responses. Transduction can be toxic (depending on auxiliaries like polybrene), number/efficiency of virus (MOI) and transduction method (spinoculation). Generally, cells need time to "calm down" and recover. Also the cell type and the overexpressed gene itself have some influence (length of bp, function, localisation ...). It can also happen that the overexpressed protein doesn't fold properly and triggers unfolded stress response what further affects cell growth and viability. So checking for transduction efficiency (e.g. FACS analysis) should be carried out at different time points until it doesn’t change much. My experience is that a GFP reporter develops rather quick (12h) while RFP needs more time (48h), yet as mentioned before, that all depends on other factors like cell line, introduced genes etc. I always check transduction efficiency after 24 hours and then by chance (e.g. when splitting the cells).







Also tagged with one or more of these keywords: transduction efficiency

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

©1999-2013 Protocol Online, All rights reserved.