Go Back   Science Forums Biology Forum Molecular Biology Forum Physics Chemistry Forum > Molecular Research Topics Forum > Cell Biology and Cell Culture > Transfection Forum
Register Search Today's Posts Mark Forums Read

Transfection Forum Transfection Forum. Cell transfection discussion including stable cells transfection, transient transfection. Troubleshooting and posting of discussions on lipofection, and reagents: Fugene, lipofectamine, and others.


Transfection of primary cells killing them

Transfection of primary cells killing them - Transfection Forum

Transfection of primary cells killing them - Transfection Forum. Cell transfection discussion including stable cells transfection, transient transfection. Troubleshooting and posting of discussions on lipofection, and reagents: Fugene, lipofectamine, and others.


Reply
 
LinkBack Thread Tools Display Modes
  #1  
Old 12-10-2009, 07:13 PM
Pipette Filler
Points: 2, Level: 1 Points: 2, Level: 1 Points: 2, Level: 1
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: Dec 2009
Posts: 1
Thanks: 0
Thanked 0 Times in 0 Posts
Default Transfection of primary cells killing them



Not sure if anyone has had this problem. We work with a primary cell line that we've been trying to transfect with pcDNA3.1. I've been using Lipofectamine 2000 to do the transfections and leave it on for 4 hours, then wash with PBS and add free media containing serum. The next day after transfection, the cells that were transfected with vector alone or vector containing my transgene are looking retracted compared to the Lipo only controls. Then when I use the cells in a spreading assay, the Lipo cells spread just fine, but the cells transfection with the vector or transgene only stick and stay round. So far we've tried using different ratios of DNA to Lipo and they seem to look better at a 1ug DNA to 1.5ul Lipo ratio, but still do not spread. Are there any other ideas for troubleshooting? My boss wants us to check the endotoxin levels of the plasmid DNA, but I'm not sure that's the problem. Help?
Reply With Quote
  #2  
Old 12-15-2009, 09:04 PM
Pipette Filler
Points: 271, Level: 5 Points: 271, Level: 5 Points: 271, Level: 5
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: Aug 2009
Posts: 5
Thanks: 0
Thanked 0 Times in 0 Posts
Default Re: Transfection of primary cells killing them

Do you like to try our transfection reagent, we can send you some sample.
Reply With Quote
Reply

Tags
cells , killing , primary , transfection


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
mouse hepatocytes isolation amicia Cell Biology and Cell Culture 39 05-11-2012 11:52 AM
Characteristics of cancerous cells aftabac Cell Biology and Cell Culture 2 08-26-2011 11:40 PM
Human Cytome Project - Update 24 Jan. 2005 Peter Van Osta Cell Biology and Cell Culture 1 08-01-2010 02:18 PM
How to isolate adult from circulation? peterish Stem Cell Forum 0 05-19-2008 10:02 PM
Human Cytome Project - Update 6 Jan. 2005 Peter Van Osta Cell Biology and Cell Culture 0 01-06-2005 11:18 AM


All times are GMT. The time now is 12:19 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 0.14834 seconds with 16 queries