I am using lentiviral vectors (2nd and 3rd generation) containing a fluorescent labeled backbone (hOSKM). After transfection of HEK239-T cells, were I could see a clear fluorescent signal after 24 hours, I put fresh viral medium directly on my target cells according to the protocol. Among several experiments, infection efficiency reflected by the fluorescent signal varied from medium to very low signal. Parallel infection with a control backbone was more succesfull in both cell lines.
(concentrations, cell densities and incubation times were stable and according to protocol)
293T cells are fluorescent due to plasmid mediated expression of the fluorescent protein. It does not mean at all that the supernatant contains functional vector.
you need to give more details on your experiment: which cells are you transducing, which fluorescent protein are you looking at and how, what range of titer do you get, what is your control vector, did you prepare it yourself, what packaging plasmids are you using .... ?
Hi Ruby,
If you need bases about relevant protocols for LV transduction, please have a look here:
https://www.geg-tech.com/Knowledge/protocols/transduction-with-lenti-one
The vector that you use (OSKM) is a very specific LV with an efficiency of production and transduction which can vary greatly, for this reason you have to optimze protocol of production and transduction. The control vector is more basic and consequently it is normal that you observe more successful transductions.
Best,
Nicolas
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