Determining viral titerviral titer
Posted 15 July 2013 - 04:39 AM
I have started with the experiments related to the transduction of Lineage negative cells with MSCV-neo vector containing my gene of interest. I am trying hard to infect the lineage negative cells but failing to do that. When I use these viruses to infect NIH3T3 cells its working fine. Please help me to determine the virus titer and also suggest some way out for my experiment.
Posted 15 July 2013 - 01:34 PM
i.e. titre (pfu/ml) = (number of plaques/dilution) x (1/volume plated(ml)).
Posted 15 July 2013 - 09:25 PM
Posted 16 July 2013 - 12:40 PM
- infect cells by your standard procedure.
- Remove inoculum.
- Mix 2% LMP agarose with 2x medium to required volume (i.e. multiply number of wells by volume) to give 1% agarose and 1x medium.
- Layer over infected cells
- Incubate for desired time (again, virus specific)
- Overlay with another layer of agarose/medium containing 0.04% neutral red (stains living cells, making plaques easier to see).
- Count plaques formed - wells of a 6 well plate with 100-500 plaques is the range to be counting. Higher than that and it gets difficult to count, lower is inaccurate.