Hello,
I typically harvest my lentivirus in the supernatant from packaging cells 48 hours after the media change from transfection. Last week, I forgot to harvest and ended up harvesting lentivirus 96 hours after media change. What will the effect be? The cells were still adhered, surprisingly. The packaging cells do express the receptor for lentiviral entry, but not to a high level. I will test the virus to determine its titer. Do you think it will it be more or less concentrated than my typical harvest? The additional time at 37C may have a negative impact on the titer, correct?
Thanks!
You will still get a good titer as cells keep producing the virus (I usually harvest the virus for several days changing the packaging culture media daily). You probably will have more viruses in this media that stayed for 96 hours (over the weekend I presume). However, the viral media may be more depleted of FBS and have more packaging cells metabolic products in it, which may be more harmful to the target cells. I would supplement it with 30-50% of fresh media before giving it to the target cells.
You will still get a good titer as cells keep producing the virus (I usually harvest the virus for several days changing the packaging culture media daily). You probably will have more viruses in this media that stayed for 96 hours (over the weekend I presume). However, the viral media may be more depleted of FBS and have more packaging cells metabolic products in it, which may be more harmful to the target cells. I would supplement it with 30-50% of fresh media before giving it to the target cells.
Hi,
I think the virus titre maybe concentrated for you which you might have to dilute and standardise. Write your observation once tested. Good wishes.
Usually, we collect the sup at 48, 72, and 96 hrs post-transfection and pool them for ultracentrifuge.
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