I produce 2nd generation lentivirus with 293T cells for T cell transduction. We concentrated our virus through 10000g centrifuge overnight. After I obtained my concentrated virus, I titrate my virus with HT1080 cells. The estimated virus titer is ranging between 8*10^7 and 2*10^8 TU/ml which said to be good enough for my continue T cell transduction. Unfortunately, we fail to got enough T cell transduction rate with these LVs( the positive rate is around 10%). The strange point is that another batch of the same Lvs which presented low titer( around 1*10^7TU/ml) could offer us much better T transduction( the positive cell rate is around 60%.
I myself got confused about the result. Can I have any suggestions to increase my T cell transduction with my Lvs? Or any cue about why these Lvs failed to transduce T cells with high transducing rate? Especially why the same Lv with low titer return much better transducing result comparing to the ones with high titer.
So many thanks!
Junjie Shao ,
Thank you for your response. We do not use polybrene. Instead, we use Transplus as transducing enhancer.
My main confusing point is that with the same transducing protocol, we got better transducing rate with lower titer-ed virus comparing the higher titer-ed virus. I personally would expect the reversed result, which said, with higher titer-ed virus, we got better transducing rate.
It is. But I am not sure what kind of toxic effect it could bring in. For the higher titer-ed and lower titer virus, we calculated and added equal amount of functional virus particles each time. The higher titer-ed virus need to add fresh medium to make the equal amount of volumes, while the lower titer-ed one didn't. In another word, we actually 'dilute' our higher titer-ed virus when we set-up the T cell transducing.
- Badges
- Science topic
- Similar topics
- Biological Science
- Microbiology
- Virology
- Virus