I am working on SF21 cell lines for a recombinant protein. During this I got several viral stock like P1, P2 and.....One most reliable method is by Plaque assay. But this is very long and time consuming process. So kindly anyone help me by telling some more rapid and reliable method with brief detail of protocol.
When faced with this problem we turned to double promoter expression plasmids. We made our protein off one promoter and a reporter gene (EGFP) off the other. Then to determine virus titers we would infect 5000 cells per well of a 96 well plate with dilutions of the virus stock. By looking at the cells through a flourescent microscope (24-36 hours later) you can determine the original virus titer just like you would from a plaque assay.
We used pBASIC promoter for EGFP and polyhedron promoter for our target genes.
Good luck.
Dear Nishat,
The above is an interesting solution. Another relatively quick method is qPCR analysis, but its major drawback is that you cannot tell whether your calculated titre reflects the amount of virions capable of infecting the host cells (some amplified viral genomes may well be derived from incompetent virions).
For that reason we stick to the limiting dilution assay (easier than the plaque assay and informative in terms of infectivity, but not really less time-consuming).
Good luck!
Radek
- Badges
- Science topic
- Similar topics
- Biological Science
- Bioscience
- Biotechnology