I need help trying to figure out why my transfections aren't working
very well. I am using NIH 3T3 cells and Lipofectin. I have tried
various ratios of Lipofectin:DNA, various times of incubation, with
Optimem or with other serum free medium. I am currently just
transfecting a reporter vector (pSVB-Gal) and then using an X-Gal assay
to identify transfected cells. Way less than 1% of the cells are
getting transfected. Everyone says these cells are so easy to transfect
and I should be able to do it blind folded. What am I doing wrong?
I start with cells that are 60 to 80% confluent and then do my assay
either 24 or 48 hours later.
Any insights would be greatly appreciated.
email: rebecca-slayton at uiowa.edu